Mixed-type

gastric

carcinomas

exhibit

more

aggressive

features

and

indicate

the

histogenesis

of

carcinomas

To

investigate

the

pathobiological

behaviors

of

gastric

mixed-type

-LRB-

MT

-RRB-

carcinomas

and

gastric

carcinogenesis

the

clinicopathological

characteristics

of

MT

carcinomas

were

analyzed

and

compared

with

intestinal-type

-LRB-

IT

-RRB-

and

diffuse-type

-LRB-

DT

-RRB-

carcinomas

The

expression

of

fragile

histine

triad

-LRB-

FHIT

-RRB-

maspin

extracellular

matrix

metalloproteinase

inducer

-LRB-

EMMPRIN

-RRB-

vascular

growth

factor

-LRB-

VEGF

-RRB-

and

E-cadherin

β-catenin

and

phosphorylated

glycogen

synthase

kinase

-LRB-

-RRB-

was

examined

on

tissue

microarrays

using

immunohistochemistry

It

was

found

that

MT

carcinomas

exhibited

large

size

deep

invasion

frequent

local

invasion

and

lymph

node

metastasis

in

comparison

with

IT

and

DT

carcinomas

-LRB-

p

-RRB-

All

the

markers

except

showed

higher

expression

in

IT

than

DT

carcinomas

-LRB-

p

-RRB-

The

expression

of

maspin

EMMPRIN

VEGF

and

membrane

E-cadherin

was

stronger

in

MT

intestinal

than

diffuse

component

-LRB-

p

-RRB-

Immunoreactivities

to

EMMPRIN

and

VEGF

were

weaker

in

IT

carcinoma

than

in

the

MT

intestinal

portion

-LRB-

p

-RRB-

while

the

opposite

was

true

for

and

-LRB-

p

-RRB-

The

MT

diffuse

component

displayed

a

higher

expression

of

FHIT

VEGF

and

than

DT

carcinoma

-LRB-

p

-RRB-

The

accumulative

survival

rate

of

the

IT

carcinoma

patients

was

higher

than

the

other

types

-LRB-

p

-RRB-

The

invasive

depth

venous

invasion

lymph

node

peritoneal

or

liver

metastasis

and

Lauren

's

classification

were

independent

prognostic

factors

for

gastric

carcinomas

-LRB-

p

-RRB-

These

findings

suggested

that

MT

carcinomas

were

also

indicated

to

be

more

aggressive

than

IT

and

DT

carcinomas

Significant

differences

were

observed

in

the

proliferation

apoptosis

angiogenesis

mucin

secretion

and

cell

adhesion

between

IT

and

DT

carcinomas

whereas

only

a

few

of

these

characteristics

showed

differences

between

the

MT

intestinal

and

diffuse

parts

thus

suggesting

that

both

the

MT

components

might

originate

from

the

stem

cells

with

similar

genetic

traits

but

follow

different

histogenic

pathways

Introduction

Gastric

carcinoma

is

the

second

leading

cause

of

cancer-related

death

behind

lung

carcinoma

despite

a

worldwide

decline

in

both

its

incidence

and

mortality

since

the

late

half

of

the

twentieth

century

-LSB-

-RSB-

It

continues

to

be

a

major

health

problem

because

of

the

slow

decrease

in

incidence

in

Asia

and

high

mortality

of

diagnosed

gastric

carcinoma

in

the

West

-LSB-

-RSB-

Generally

the

prognosis

of

patients

with

gastric

carcinoma

is

somewhat

dependent

on

its

histological

type

such

as

intestinal

type

-LRB-

IT

-RRB-

or

diffuse

type

-LRB-

DT

-RRB-

as

defined

by

Lauren

-LSB-

-RSB-

Although

gastric

carcinoma

is

a

malignant

tumor

originating

from

the

same

gastric

epithelium

its

morphological

features

vary

substantially

among

individual

patients

Histologically

IT

carcinoma

principally

includes

papillary

well-differentiated

and

moderately-differentiated

or

mucinous

adenocarcinoma

without

signet

ring

carcinoma

-LRB-

SRC

-RRB-

cells

whereas

DT

mainly

consists

of

the

SRC

poorly

differentiated

and

undifferentiated

adenocarcinoma

of

WHO

classification

-LSB-

-RSB-

Although

Kushima

and

Hattori

-LSB-

-RSB-

firstly

proposed

an

entity

of

gastric-type

carcinoma

and

terminologically

seemed

homologous

to

the

IT

one

the

carcinoma

cells

of

this

type

closely

resemble

normal

foveolar

cells

on

the

basis

of

tissue

morphology

and

mucin

properties

and

are

considered

to

have

been

derived

from

foveolar

hyperplasia

or

pyloric

gland

adenoma

-LSB-

-RSB-

Recently

our

group

also

focused

on

Lauren

's

classification

and

found

that

IT

carcinoma

frequently

occurred

in

old

men

while

DT

was

more

frequent

in

comparatively

young

women

The

latter

group

was

more

inclined

to

invasion

into

the

muscularis

propria

lymphatic

invasion

and

lymph

node

metastasis

and

belonged

to

a

higher

TNM

staging

in

comparison

to

its

IT

counterparts

Further

analysis

demonstrated

that

IT

gastric

carcinomas

with

a

more

favorable

prognosis

were

prone

to

high

levels

of

proliferation

and

apoptosis

and

also

always

accompanied

by

a

strong

expression

of

the

fragile

histine

triad

-LRB-

FHIT

-RRB-

phosphatase

and

tensin

homology

deleted

from

human

chromosome

-LRB-

PTEN

-RRB-

and

extracellular

matrix

metalloproteinase

inducer

-LRB-

EMMPRIN

-RRB-

-LSB-

-RSB-

Additionally

many

genetic

and

epigenetic

changes

such

as

mutation

deletion

loss

of

heterozygosity

methylation

and

microsatellite

instability

were

differentially

observed

in

both

histological

types

of

gastric

carcinoma

-LSB-

-RSB-

These

data

indicate

that

different

carcinogenic

pathways

exist

for

gastric

IT

and

DT

carcinomas

Although

carcinogenesis

is

a

multistage

process

which

consists

of

a

multi-factorial

etiology

resulting

from

gene

environment

interaction

the

intestinal

type

is

frequently

related

to

environmental

factors

like

diet

and

predominates

in

areas

with

a

high

incidence

of

this

disease

In

contrast

the

diffuse

type

is

thought

to

be

of

genetic

origin

and

evenly

distributed

worldwide

-LSB-

-RSB-

It

is

commonly

believed

that

Lauren

's

classification

is

valuable

for

both

the

epidemiological

studies

and

gastric

carcinogenesis

However

a

small

number

of

gastric

carcinomas

remain

characterized

as

unclassified

or

mixed-type

-LRB-

MT

-RRB-

ones

which

are

comprised

histologically

of

non-homogenous

mixtures

of

IT

and

DT

carcinomas

Stelzner

and

Emmrich

-LSB-

-RSB-

found

that

MT

carcinomas

showed

a

deep

infiltration

of

the

gastric

wall

frequent

regional

lymph

node

metastasis

and

high

staging

in

comparison

to

the

other

two

types

Machado

et

al.

-LSB-

-RSB-

reported

that

inactivating

E-cadherin

mutations

were

exclusively

observed

in

the

diffuse

component

of

the

tumors

thus

suggesting

that

MT

carcinomas

displayed

phenotypic

divergence

A

recent

study

showed

that

bone-marrow-derived

cells

-LRB-

BMDCs

-RRB-

could

progress

through

metaplasia

and

dysplasia

to

intraepithelial

cancer

under

the

induction

of

the

carcinogen

Helicobacter

pylori

-LSB-

-RSB-

Therefore

the

analysis

of

distinct

MT

carcinoma

components

can

be

of

remarkable

pathogenic

significance

if

it

would

be

true

that

carcinoma

cells

originate

from

BMDCs

In

the

present

study

cases

of

gastric

carcinomas

were

collected

to

further

clarify

the

pathological

characteristics

of

MT

carcinoma

and

the

gastric

carcinogenesis

pathways

using

a

combination

of

tissue

microarray

-LRB-

TMA

-RRB-

and

immunohistochemical

techniques

Materials

and

methods

Subjects

A

total

of

gastric

carcinomas

were

collected

from

surgical

resection

in

Kouseiren

Takaoka

Hospital

between

and

The

patients

with

carcinomas

included

men

and

women

-LRB-

years

mean

years

-RRB-

Among

them

cases

were

demonstrated

with

lymph

node

metastasis

and

with

liver

metastasis

of

these

cases

underwent

either

chemotherapy

or

radiotherapy

before

surgery

All

patients

gave

their

informed

consent

for

the

use

of

tumor

tissue

specimens

for

clinical

research

and

the

University

Ethical

Committee

approved

the

research

protocol

All

patients

were

followed

up

by

consulting

their

case

documents

and

through

telephone

interviews

Pathology

All

tissue

specimens

were

fixed

in

neutralized

formaldehyde

embedded

in

paraffin

and

cut

into

sections

These

sections

were

stained

by

hematoxylin

and

eosin

-LRB-

HE

-RRB-

to

confirm

their

histological

diagnosis

and

other

microscopic

characteristics

The

staging

for

each

gastric

carcinoma

was

evaluated

according

to

the

TNM

system

of

the

Internationale

Contre

le

Cancer

-LRB-

UICC

-RRB-

indicating

the

extent

of

tumor

spread

-LSB-

-RSB-

The

histomorphological

architecture

of

the

tumors

was

expressed

according

to

Lauren

's

classification

-LSB-

-RSB-

The

growth

patterns

were

divided

into

five

groups

based

on

a

modification

of

Emmrich

's

method

-LSB-

-RSB-

Briefly

group

I

showed

a

combination

of

two

components

with

diffuse

distribution

Group

II

displayed

both

components

with

the

border

clearly

visible

Group

III

represented

some

signet

cells

in

the

intestinal

and

diffuse

components

Group

IV

were

mucinous

carcinomas

with

signet

cells

Group

V

exhibited

separately

intestinal

and

diffuse

components

in

the

gastric

wall

In

addition

the

depth

of

invasion

lymphatic

and

venous

invasion

and

peritoneal

dissemination

were

all

determined

Tissue

microarray

IT

and

DT

carcinomas

as

well

as

both

components

of

the

mixed

type

were

identified

in

HE

stained

sections

of

the

selected

tumor

cases

and

a

tissue

core

of

each

donor

block

was

punched

out

and

transferred

to

a

recipient

block

with

a

maximum

of

cores

using

a

Tissue

Microarrayer

-LRB-

AZUMAYA

Tokyo

Japan

-RRB-

Four-micrometer-thick

sections

were

cut

from

the

recipient

block

and

transferred

to

poly-lysine-coated

glass

slides

HE

staining

was

performed

on

TMA

for

confirmation

of

the

tumor

tissue

-LRB-

Fig.

-RRB-

Fig.

staining

of

TMA

of

gastric

carcinomas

Immunohistochemistry

Consecutive

sections

were

deparaffinized

with

xylene

dehydrated

with

alcohol

and

subjected

to

antigen

retrieval

by

irradiating

in

target

retrieval

solution

-LRB-

TRS

DAKO

Carpinteria

UAS

-RRB-

for

min

in

a

microwave

oven

-LRB-

Oriental

rotor

Lmt

Co.

Tokyo

Japan

-RRB-

Five

percent

bovine

serum

albumin

-LRB-

BSA

-RRB-

was

then

applied

for

min

to

prevent

non-specific

binding

The

sections

were

incubated

with

primary

antibodies

for

min

then

treated

with

the

anti-mouse

or

anti-rabbit

Envison-PO

-LRB-

DAKO

USA

-RRB-

antibodies

for

min

All

incubations

were

performed

in

the

microwave

oven

for

intermittent

irradiation

as

described

previously

-LSB-

-RSB-

After

each

treatment

the

slides

were

washed

with

TBST

-LRB-

mM

Tris

HCl

mM

NaCl

Tween

-RRB-

three

times

for

min

The

primary

antibodies

are

summarized

in

Table

All

immunostaining

was

visualized

with

diaminobenzidine

-LRB-

DAB

-RRB-

with

reaction

and

counterstained

with

Mayer

's

hematoxylin

Omission

of

the

primary

antibody

was

used

as

a

negative

control

Table

Primary

antibodies

used

in

this

Carpinteria

Fremont

Newcastle

upon

Tyne

UKRead-to-useEMMPRINMouseNovocastra

Fremont

Otsu

CA

The

immunoreactivity

to

and

was

localized

in

the

nucleus

FHIT

vascular

growth

factor

-LRB-

VEGF

-RRB-

maspin

phosphorylated

glycogen

synthase

kinase

-LRB-

-RRB-

and

in

the

cytoplasm

EMMPRIN

and

in

the

cytoplasm

and

membrane

and

E-cadherin

in

the

membrane

and

β-catenin

in

the

nucleus

cytoplasm

and

membrane

-LRB-

Fig.

-RRB-

All

evaluations

were

performed

blindly

by

two

independent

observers

-LRB-

Takano

Y

and

Zheng

HC

-RRB-

Fig.

of

TMA

of

gastric

carcinomas

The

immunoreactivity

to

-LRB-

a

-RRB-

and

-LRB-

c

-RRB-

was

localized

in

the

nucleus

FHIT

-LRB-

d

-RRB-

-LRB-

b

-RRB-

VEGF

-LRB-

g

-RRB-

maspin

-LRB-

e

-RRB-

-LRB-

h

-RRB-

-LRB-

i

-RRB-

-LRB-

k

-RRB-

-LRB-

l

-RRB-

were

seen

in

the

cytoplasm

EMMPRIN

-LRB-

f

-RRB-

and

-LRB-

j

-RRB-

were

seen

in

the

cytoplasm

and

the

plasma

membrane

-LRB-

m

-RRB-

and

E-cadherin

-LRB-

n

-RRB-

in

the

plasma

membrane

and

β-catenin

-LRB-

o

-RRB-

in

the

nucleus

cytoplasm

and

plasma

membrane

These

molecules

were

strongly

expressed

in

the

intestinal-type

gastric

carcinomas

Statistical

analysis

The

statistical

evaluation

was

performed

using

Fisher

's

exact

possibility

to

differentiate

the

rates

and

the

Mann

Whitney

U

test

to

differentiate

the

means

of

the

different

groups

Kaplan

Meier

survival

plots

were

generated

and

comparisons

between

the

survival

curves

were

made

with

the

log-rank

statistic

The

Cox

proportional

hazards

model

was

employed

for

multivariate

analysis

p

was

considered

to

represent

a

statistically

significant

difference

The

SPSS

software

program

was

employed

to

analyze

all

data

Results

Clinicopathological

characteristics

of

gastric

IT

DT

and

MT

carcinomas

As

summarized

in

Table

IT

cases

were

identified

-LRB-

-RRB-

DT

-LRB-

-RRB-

and

MT

-LRB-

-RRB-

among

the

gastric

carcinomas

in

this

study

Patients

with

IT

carcinoma

were

found

to

be

significantly

older

than

those

with

DT

-LRB-

p

-RRB-

The

male

and

female

ratios

in

the

carcinoma

cases

gradually

decreased

from

IT

to

DT

through

the

MT

groups

-LRB-

p

-RRB-

It

was

the

same

for

lower

TNM

staging

in

the

three

groups

-LRB-

p

-RRB-

The

positive

rate

of

peritoneal

spread

was

higher

in

the

DT

and

MT

cases

than

IT

-LRB-

p

-RRB-

MT

carcinomas

showed

aggressive

characteristics

such

as

large

size

deep

invasion

frequent

local

invasion

and

lymph

node

metastasis

compared

to

the

IT

and

DT

-LRB-

p

-RRB-

There

was

no

significance

between

the

three

types

of

gastric

carcinomas

in

the

incidence

of

the

liver

metastasis

-LRB-

p

-RRB-

Table

Comparison

of

the

clinicopathological

features

in

gastric

IT

DT

and

MT

carcinomasClinicopathological

featuresIntestinal-type

carcinomaDiffuse-type

carcinomaMixed-type

carcinomaCase

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

Age

-LRB-

mean

SD

years

-RRB-

Sex

-LRB-

male

female

-RRB-

Tumor

size

-LRB-

mean

SD

cm

-RRB-

Invasion

into

muscularis

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

Lymphatic

invasion

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

Venous

invasion

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

Lymph

node

metastasis

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

Liver

metastasis

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

Peritoneal

spread

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

TNM

staging

-LRB-

O

I

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

p

compared

with

intestinal

and

mixed-type

carcinomas

p

compared

with

intestinal

and

diffuse-type

carcinomas

with

both

analyzed

p

compared

with

diffuse

and

mixed-type

carcinomas

Pathological

behaviors

of

gastric

MT

carcinomas

with

different

growth

patterns

or

histological

distribution

of

components

According

to

the

growth

pattern

the

intestinal

and

diffuse

components

diffusely

distributed

without

absolute

border

in

group

I

of

the

MT

carcinomas

which

occurred

more

frequently

-LRB-

-RRB-

than

the

other

four

groups

As

indicated

in

Table

most

groups

III

and

V

gastric

carcinomas

were

generally

of

early

stage

with

less

local

invasion

infrequent

lymph

node

metastasis

and

low

TNM

staging

In

this

study

only

four

advanced

gastric

carcinomas

belonged

to

group

IV

among

which

three

cases

were

women

and

all

were

accompanied

with

lymphangiogenic

invasion

lymph

node

metastasis

and

high

TNM

staging

Although

the

main

histological

component

was

different

from

that

in

lymph

node

metastasis

the

statistical

data

revealed

that

there

was

histological

consistency

between

primary

and

lymph

node

metastasis

foci

-LRB-

p

-RRB-

There

were

no

remarkable

differences

in

the

pathological

behaviors

of

MT

carcinomas

with

different

ratios

of

intestinal

and

diffuse

portions

-LRB-

p

-RRB-

Table

Pathological

behaviors

of

gastric

MT

carcinomasClinicopathological

featuresnSex

-LRB-

male

-RRB-

Invasion

into

MPLymphatic

invasionVenous

invasionLymph

node

metastasisStagingnI

DI

DI

D

-LRB-

O

I

-RRB-

Growth

pattern

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

Histological

appearance

Intestinal

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

Intestinal

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

Intestinal

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

MP

Muscularis

propria

in

lymph

node

metastatic

foci

I

D

intestinal

diffuse

I

D

intestinal

diffuse

I

=D

intestinal

diffuse

Immunohistochemical

analysis

of

intestinal

or

diffuse

components

from

three

types

of

gastric

carcinomas

Table

demonstrates

that

all

the

markers

except

were

expressed

at

higher

levels

in

IT

carcinomas

than

in

MT

ones

-LRB-

p

-RRB-

The

expression

of

maspin

EMMPRIN

VEGF

and

E-cadherin

was

stronger

in

the

intestinal

component

of

MT

carcinomas

than

in

their

diffuse

counterpart

-LRB-

p

-RRB-

Immunoreactivities

to

EMMPRIN

and

VEGF

were

weaker

in

the

intestinal-type

carcinomas

than

in

the

intestinal

component

of

the

MT

-LRB-

p

-RRB-

while

it

was

the

opposite

for

and

-LRB-

p

-RRB-

In

contrast

the

diffuse

components

of

the

MT

carcinomas

showed

greater

expression

of

FHIT

VEGF

and

than

DT

carcinomas

-LRB-

p

-RRB-

Table

Immunohistochemical

analysis

in

gastric

IT

DT

and

MT

carcinomasBiological

markersIntestinal-type

carcinomaMixed-type

carcinomaDiffuse-type

carcinomaIntestinal

partDiffuse

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

Membrane

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

p

compared

with

the

diffuse-type

carcinomas

p

compared

with

intestinal-part

carcinomas

p

compared

with

diffuse-part

carcinomas

Patients

outcome

with

different

gastric

carcinomas

Follow-up

information

for

carcinoma

patients

was

used

before

for

a

period

ranging

from

days

to

years

-LRB-

mean

months

-RRB-

Figure

shows

the

survival

curves

stratified

according

to

Lauren

's

classification

Kaplan

Meier

analysis

indicated

that

the

patients

with

IT

carcinoma

had

a

higher

cumulative

survival

rate

than

those

with

the

DT

and

MT

lesions

-LRB-

p

-RRB-

Although

the

MT

carcinoma

patients

survival

rate

was

comparatively

lower

than

the

DT

ones

there

was

no

statistical

significance

-LRB-

p

-RRB-

Multivariate

analysis

demonstrated

that

invasive

depth

venous

invasion

lymph

node

peritoneal

or

liver

metastasis

and

Lauren

's

classification

were

independent

factors

for

the

poor

prognosis

of

gastric

carcinoma

patients

-LRB-

p

Table

-RRB-

Fig.

between

prognosis

of

carcinoma

patients

and

Lauren

's

classification

Kaplan

Meier

curves

of

cumulative

survival

rate

for

the

patients

with

gastric

carcinoma

according

to

Lauren

's

classification

IT

Intestinal

type

DT

diffuse

type

MT

mixed

typeTable

Multivariate

analysis

of

the

clinicopathological

variables

of

gastric

carcinomasClinicopathological

parametersRelative

risk

-LRB-

CI

-RRB-

p

ValueAge

-LRB-

years

-RRB-

-LRB-

-RRB-

Sex

-LRB-

male

female

-RRB-

-LRB-

-RRB-

Tumor

size

-LRB-

cm

-RRB-

-LRB-

-RRB-

Invasive

depth

-LRB-

Tis

-RRB-

-LRB-

-RRB-

Lymphatic

invasion

-LRB-

-RRB-

-LRB-

-RRB-

Venous

invasion

-LRB-

-RRB-

-LRB-

-RRB-

Lymph

node

metastasis

-LRB-

-RRB-

-LRB-

-RRB-

Peritoneal

spread

-LRB-

-RRB-

-LRB-

-RRB-

Liver

metastasis

-LRB-

-RRB-

-LRB-

-RRB-

Lauren

's

classification

-LRB-

intestinal/diffuse/mixed

-RRB-

-LRB-

-RRB-

CI

Confidence

interval

Discussion

Histologically

Lauren

's

classification

of

gastric

carcinomas

is

based

on

the

morphological

appearances

IT

carcinomas

are

characterized

by

cohesive

carcinoma

cells

forming

gland-like

tubular

structures

with

an

expanding

or

infiltrative

growth

pattern

like

a

papillary

and

well-differentiated

adenocarcinoma

However

the

cell

cohesion

is

less

apparent

or

absent

in

DT

carcinomas

and

cancer

cells

diffusely

spread

in

the

gastric

wall

as

poorly

differentiated

adenocarcinomas

SRC

or

undifferentiated

carcinomas

-LSB-

-RSB-

However

there

are

two

types

of

histological

distribution

in

the

MT

carcinoma

as

described

by

Lauren

-LSB-

-RSB-

In

this

study

carcinoma

diagnosed

as

MT

made

up

of

the

whole

series

in

this

population

more

than

that

reported

by

Borch

et

al.

-LRB-

-RRB-

Lauren

-LRB-

-RRB-

and

Stelzner

and

Emmrich

-LRB-

-RRB-

and

less

than

that

reported

by

Carneiro

et

al.

-LRB-

-RRB-

-LSB-

-RSB-

This

discrepancy

might

be

due

to

the

differences

in

diagnosis

criteria

geographical

regions

human

races

living

habits

and

so

forth

The

age

and

sex

distribution

of

the

patients

with

MT

carcinoma

was

intermediate

although

IT

carcinomas

frequently

occurred

in

old

men

compared

with

DT

carcinomas

Morphologically

there

are

both

intestinal

and

diffuse

components

in

MT

carcinomas

and

they

were

found

to

be

significantly

larger

more

advanced

and

more

frequently

spread

into

local

lymphatic

and

venous

vessels

or

regional

lymph

nodes

compared

with

IT

and

DT

carcinomas

in

the

present

study

consistent

with

other

reports

-LSB-

-RSB-

Kozuki

et

al.

-LSB-

-RSB-

found

that

prominent

lymphatic

permeation

and

lymph

node

metastasis

were

more

frequently

observed

in

MT

than

in

the

pure

type

of

gastric

carcinomas

It

was

previously

documented

that

the

MT

carcinomas

showed

a

deeper

infiltration

of

the

gastric

wall

a

higher

metastatic

rate

to

the

regional

lymph

nodes

and

the

need

for

higher

staging

with

the

TNM

system

of

UICC

classification

in

contrast

to

IT

and

MT

carcinomas

-LSB-

-RSB-

These

findings

suggest

that

the

pathobiological

characteristics

of

the

MT

carcinomas

were

more

aggressive

than

the

other

two

types

which

accounted

for

their

comparatively

poor

prognosis

observed

in

the

present

study

although

there

was

no

statistical

significance

Among

the

MT

carcinomas

five

groups

were

classified

according

to

their

growth

pattern

Half

of

the

present

cases

belonged

to

group

I

which

means

there

was

diffuse

distribution

of

the

intestinal

and

diffuse

component

without

clear

borders

in

most

of

the

MT

cases

In

general

groups

III

and

IV

of

MT

carcinomas

did

not

exhibit

deep

invasion

comparatively

high

staging

or

frequent

spreading

compared

with

other

types

However

group

IV

of

carcinomas

with

mucin

and

signet

cells

was

more

advanced

with

wide

spreading

including

local

vessels

and

lymph

node

metastasis

Therefore

surgeons

should

carefully

treat

the

patients

with

MT

carcinomas

in

clinical

practice

because

of

their

aggressive

characteristics

and

poor

prognosis

especially

those

classified

as

group

IV

Furthermore

it

was

found

that

the

major

component

in

the

MT

primary

foci

morphologically

paralleled

to

that

in

the

corresponding

lymph

node

metastasis

although

the

converse

situation

also

appeared

This

inconsistent

phenomenon

could

be

explained

by

sampling

bias

or

the

metastatic

potential

of

individual

carcinoma

cells

Compared

with

IT

carcinomas

the

MT

counterpart

always

exhibited

more

severe

characteristics

including

invasion

into

muscularis

propria

lymphatic

invasion

and

lymph

node

metastasis

and

high

TNM

staging

in

the

present

study

as

observed

by

Lauren

and

other

studies

-LSB-

-RSB-

To

clarify

the

pheno/genotypes

of

gastric

carcinoma

the

expression

of

FHIT

maspin

VEGF

EMMPRIN

E-cadherin

and

β-catenin

was

examined

by

immunostaining

FHIT

and

maspin

as

tumor

suppressor

genes

play

important

roles

in

regulating

the

balance

between

the

proliferation

and

apoptosis

of

cancer

cells

-LSB-

-RSB-

antigen

is

present

in

the

nuclei

of

cells

undergoing

proliferation

and

should

be

regarded

as

a

good

marker

for

cell

proliferation

-LSB-

-RSB-

is

responsible

for

the

cleavage

of

poly

-LRB-

ADP-ribose

-RRB-

polymerase

actin

and

sterol

regulatory

element

binding

protein

and

reflects

the

apoptotic

level

as

a

key

protease

in

the

cascade

reaction

of

the

apoptotic

pathway

-LSB-

-RSB-

In

vivo

and

vitro

evidences

indicated

that

EMMPRIN

and

VEGF

are

involved

in

angiogenic

processes

in

malignancies

-LSB-

-RSB-

Glycogen

synthase

-LRB-

-RRB-

belongs

to

the

serine/threonine

protein

kinase

family

and

is

also

involved

in

regulating

the

balance

between

proliferation

and

apoptosis

and

can

be

inactivated

via

phosphorylation

by

kinase

Akt

certain

isoforms

of

proteins

kinase

and

cyclic

AMP-dependent

protein

kinase

-LSB-

-RSB-

and

are

markers

for

intestinal

goblet

cells

superficial

epithelium

and

gastric

pyloric

gland

cells

respectively

which

can

reflect

the

mucin

secretion

and

variations

in

gastrointestinal

malignancies

is

a

heterodimeric

glycoprotein

complex

and

expressed

in

several

human

epithelial

carcinomas

-LSB-

-RSB-

is

a

cell

surface

glycoprotein

involved

in

cell/cell

and

cell/matrix

interactions

overexpression

has

been

linked

to

the

growth

and

spread

of

a

range

of

different

types

of

malignancies

-LSB-

-RSB-

The

E-cadherin

can

interact

with

β-catenin

to

form

a

complex

which

is

closely

linked

to

cell

adhesion

and

differentiation

-LSB-

-RSB-

In

the

present

study

all

of

the

markers

except

showed

greater

expression

in

the

IT

than

in

the

DT

carcinomas

Although

some

evidences

indicate

that

was

strongly

expressed

in

gastric

DT

carcinoma

in

comparison

with

IT

ones

there

was

no

statistical

difference

in

line

with

our

finding

-LSB-

-RSB-

These

data

suggested

that

there

were

significant

differences

in

the

proliferation

apoptosis

angiogenesis

mucin

production

and

cell

adhesion

between

the

IT

and

DT

carcinomas

and

these

molecules

mechanistically

contributed

to

the

molecular

distinction

in

the

morphological

behavioral

and

histogenic

aspects

between

both

types

of

gastric

carcinomas

In

general

DT

carcinoma

is

believed

to

derive

de

novo

from

the

peripheral

stem

cells

of

gastric

gland

neck

proliferative

zone

without

a

recognizable

precursor

lesion

except

hereditary

diffuse

gastric

cancer

because

in

situ

carcinoma

or

globoid

dysplasia

is

its

precancerous

lesion

respectively

according

to

the

Chinese

and

Western

pathologists

opinions

-LSB-

-RSB-

When

long-standing

gastric

inflammation

causes

tissue

injury

and

stem

cell

failure

with

time

BMDCs

are

recruited

and

engrafted

into

the

tissue

stem

cell

niche

where

BMDCs

can

behave

in

a

way

indistinguishable

from

endogenous

tissue

stem

cells

With

continued

inflammation

and

injury

they

can

undergo

sustained

proliferation

and

malignant

transformation

into

IT

carcinomas

passing

through

precancerous

stages

of

metaplasia

and

dysplasia

when

genetic

defects

such

as

mutation

deletion

or

rearrangement

are

accumulated

resulting

in

corruption

of

the

balance

between

proliferation

and

apoptosis

-LSB-

-RSB-

Likewise

dedifferentiation

of

intestinal

to

diffuse

carcinoma

had

been

identified

as

another

histogenic

pathway

according

to

the

histological

heterogeneity

of

tumor

cells

especially

in

the

carcinomas

containing

moderately

and

poorly

differentiated

components

with

similar

morphological

appearance

and

diffuse

distribution

If

so

it

is

possible

that

the

distinct

components

in

MT

carcinomas

arise

from

the

stem

cells

with

common

genetic

traits

and

follow

different

carcinogenic

pathways

Conversely

diffuse

carcinoma

could

be

derived

from

heterogeneity

of

the

intestinal

counterpart

According

to

our

present

data

no

differences

in

the

expression

of

FHIT

and

membrane

β-catenin

were

observed

between

the

intestinal

and

diffuse

components

of

MT

carcinomas

which

supports

the

possibility

of

similar

origin

or

dedifferentiation

However

increased

expression

of

EMMPRIN

VEGF

and

E-cadherin

in

the

intestinal

component

compared

to

the

diffuse

counterpart

also

provided

evidence

that

the

original

carcinoma

cells

might

undergo

distinct

carcinogenic

routes

resulting

in

the

morphological

distinction

of

both

components

Both

the

intestinal

and

diffuse

components

in

MT

carcinomas

had

increased

expression

of

EMMPRIN

and

VEGF

and

reduced

E-cadherin

Serum

VEGF

levels

were

found

to

be

significantly

higher

in

patients

with

MT

gastric

carcinomas

than

those

with

pure

lesions

-LSB-

-RSB-

It

was

proposed

that

MT

carcinomas

biologically

displayed

more

aggressive

behaviors

than

other

types

including

decreased

cell

adhesion

increased

proliferation

and

angiogenesis

Most

of

the

evidences

in

the

present

data

indicated

that

the

pathological

behaviors

of

MT

carcinomas

were

more

aggressive

than

the

other

types

which

was

closely

linked

to

the

prognosis

It

was

previously

documented

that

the

MT

patients

survival

is

significantly

worse

than

those

with

IT

or

DT

carcinomas

regardless

of

their

location

-LSB-

-RSB-

However

this

study

demonstrated

that

there

was

no

significance

between

the

MT

and

DT

carcinoma

patients

survival

rates

although

the

former

was

lower

than

the

latter

Additionally

both

survival

rates

were

lower

than

that

with

IT

carcinomas

To

avoid

sampling

bias

from

a

shorter

follow-up

time

the

older

cases

were

chosen

for

the

survival

analysis

but

they

yielded

consistent

results

Furthermore

Cox

's

hazard

proportional

analysis

indicated

that

the

invasive

depth

venous

invasion

lymph

node

peritoneal

or

liver

metastasis

and

Lauren

's

classification

were

independent

prognostic

factors

for

gastric

carcinomas

In

summary

IT

carcinoma

which

is

positively

correlated

with

favorable

prognosis

frequently

displayed

high

levels

of

proliferation

apoptosis

angiogenesis

mucin

production

and

cell

adhesion

Gastric

MT

carcinoma

showed

more

aggressive

behaviors

than

IT

and

DT

ones

There

was

a

significant

difference

in

the

proliferation

apoptosis

angiogenesis

mucin

secretion

and

cell

adhesion

between

the

IT

and

DT

carcinomas

whereas

only

a

few

characteristics

were

differentially

detected

in

the

intestinal

and

diffuse

component

of

the

mixed-type

carcinoma

suggesting

that

different

components

of

MT

carcinoma

might

originate

from

common

stem

cells

but

follow

distinct

histogenic

pathways

Furthermore

these

results

confirm

that

Lauren

's

classification

is

significant

regarding

the

histopathogenesis

and

differentiation

and

considered

as

a

guide

to

the

clinical

treatment

of

gastric

carcinoma

O O O O O O O O O O O O O O O O B I O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O

gastric carcinoma
mixed type
pathobiological behaviors
carcinogenesis


The effects of laparoscopic cholecystectomy , hysterectomy , and appendectomy on nosocomial infection risks Background Recent reviews of the literature have concluded that additional , well-defined studies are required to clarify the superiority of laparoscopic or open surgery . This paper presents precise estimates of nosocomial infection risks associated with laparoscopic as compared to open surgery in three procedures : cholecystectomy , appendectomy , and hysterectomy . With the realization of smaller incisions , better cosmesis , less postoperative pain , same-day surgery , speedier postoperative recovery , and the potential for reduced complications , laparoscopic approaches have all but replaced the traditional laparotomic alternatives for certain commonly performed surgical procedures . However , the widespread adoption of laparoscopic techniques into the overall surgical armamentarium has been slowed by a variety of factors , including the learning curves required to integrate new levels of depth perception and fine dexterity , longer operating times without commensurate economic reward , and the nullification of savings from earlier hospital discharge by the cost of disposable surgical instrumentation . Nevertheless , the laparoscopic approach is now widely accepted as the gold standard for cholecystectomy and the surgical treatment of gastroesophageal reflux -LSB- 1 , 2 -RSB- . Laparoscopic appendectomy has been controversial since its introduction in the early 1980s , particularly for cases of complicated appendicitis . Although laparoscopic appendectomies now account for almost half of appendectomies in the United States -LSB- 3 -RSB- , several recent reviews of the literature have concluded that additional , well-defined studies will be required to clarify the superiority of laparoscopic or open approaches -LSB- 4 -- 6 -RSB- . Despite the demonstration that abdominal hysterectomy is associated with higher morbidity and worse outcomes when compared to the vaginal or laparoscopic approach , the majority of hysterectomies worldwide are still performed in this fashion . For the remaining cases , laparoscopic hysterectomy is least apt to be performed -LSB- 7 , 8 -RSB- . Although laparoscopy facilitates vaginal hysterectomy for the larger uterus , allows for the concurrent diagnosis and treatment of benign pelvic conditions such as endometriosis or pelvic adhesions , permits concomitant adnexal surgery , and provides the ability to secure and reaffirm intraperitoneal hemostasis at the end of the procedure -LSB- 9 -RSB- , a meta-analysis of randomized controlled trials comparing different types of hysterectomy published by the Cochrane Collaboration failed to clearly demonstrate the superiority of laparoscopic hysterectomy over vaginal hysterectomy -LSB- 10 -RSB- . Some studies have reported that laparoscopic approaches are associated with lower risks of surgical site infections than their open counterparts -LSB- 11 -- 13 -RSB- , but the effects of laparoscopic surgery on overall nosocomial -LRB- hospital-acquired -RRB- infection risks have not been established . Since a significant number of nosocomial infections in surgical patients occur after discharge -LSB- 14 , 15 -RSB- , it is likely that comparisons of laparoscopic and open surgeries have underestimated risks . Nosocomial infections are a leading cause of death in the United States , affecting two to three million patients annually -LSB- 16 -RSB- . Starting in 2009 , Centers for Medicare and Medicaid Services -LRB- CMS -RRB- will stop reimbursing hospitals for certain complications including surgical-site infections , catheter-associated urinary tract infections , and central-line associated bloodstream infections -LSB- 17 -RSB- . Therefore , the establishment of nosocomial infection risks in general is important . The objective of this study is to obtain more precise estimates of nosocomial infection risks associated with laparoscopic and open approaches for cholecystectomy , appendectomy , and hysterectomy . We hypothesize that laparoscopic surgery will reduce the risk of nosocomial infections for each of these surgical modalities . To test these hypotheses we performed a retrospective analysis of more than 11,000 admissions , each with one of the procedures of interest , from 22 hospitals that have implemented a nosocomial infection monitoring system that can detect nosocomial infections up to 30 days post discharge . Methods The Nosocomial Infection Marker -LRB- NIM -RRB- The Nosocomial Infection Marker -LRB- NIM , patent pending , Cardinal Health -RRB- monitors and tracks nosocomial infection rates for hospitals and communities . Cardinal Health extracts data from client facilities on an ongoing basis using a secure , Health Insurance Portability and Accoutability Act - -LRB- HIPAA -RRB- compliant method . Data are cleaned and mapped in real time as they arrive at the Cardinal Health data center by proprietary software systems . Rare exceptions that are not electronically modeled are modeled by technical and clinical experts , processed and loaded . The new models are then used by the systems to process like data in the future . The Nosocomial Infection Marker is a computer algorithm that identifies the existence of nosocomial infections at the microbiological level . Specifically , the NIM algorithm distinguishes likely pathogens from contaminants , identifies duplicate isolates , and temporally determines hospital versus community-acquired pathogen acquisition -LSB- 18 -RSB- . In a multihospital study using comprehensive medical records review and gold-standard infectious disease physician discrepancy resolution , the NIM algorithm identified nosocomial infections with 86 % sensitivity and 98.5 % specificity hospital-wide , statistically outperforming Centers for Disease Control -LRB- CDC -RRB- case finding methods -LSB- 18 -RSB- . Unlike the NIM , traditional CDC case finding methods are subjectively applied with inconsistent results and are only used for certain types of infections , mostly in ICUs -LSB- 18 , 19 -RSB- . Like the NIM , the performance characteristics of CDC methods have only been formally evaluated in one study -LSB- 19 -RSB- . Data The Nosocomial Infection Marker -LRB- NIM -RRB- was used to identify nosocomial infections during hospitalization and post discharge . Data were extracted from the Cardinal Health -LRB- CAH -RRB- data repository for the period September 1 , 2004 through December 31 , 2006 from 22 hospitals in 15 states . Hospitals with matching International Statistical Classification of Diseases and Related Health Problems -LRB- ICD-9 -RRB- procedure codes and more than 100 admissions with a primary ICD-9 procedure for cholecystectomy , appendectomy , or hysterectomy were eligible . These hospitals had a median number of beds of 359 , with an interquartile range from 191 to 483 beds ; one hospital exceeded 1,000 beds and two had fewer than 150 . Mean hospital CMI was 1.56 with a standard deviation of 0.22 . Admissions with primary Diagnosis Related Group -LRB- DRGs -RRB- listed in Table 1 comprised more than 95 % of eligible admissions . When these admissions were restricted to adults 18 years and older for cholecystectomy and hysterectomy , and patients 2 years and older for appendectomy , 11,662 admissions were available for analysis . Table 1DRGs included in the analysis % admissions NIM rate % laparoscopySimple presentations -LRB- complexity = 0 -RRB- 166Appendectomy W/O complicated Principal Diag W Cc3 .162.7167.21167 Appendectomy W/O Complicated Principal Diag W/O Cc14 .531.1271.66195 Cholecystectomy W C.D.E. W Cc0 .5214.7516.39196 Cholecystectomy W C.D.E. W/O Cc0 .210.0025.00197 Cholecystectomy Except By Laparoscope W/O C.D.E. W Cc3 .649.4311.79198 Cholecystectomy Except By Laparoscope W/O C.D.E. W/O Cc2 .012.5640.60358 Uterine & Adnexa Proc For Non-Malignancy W Cc11 .433.3036.53359 Uterine & Adnexa Proc For Non-Malignancy W/O Cc25 .721.2047.92493 Laparoscopic Cholecystectomy W/O C.D.E. W Cc14 .864.15100494 Laparoscopic Cholecystectomy W/O C.D.E. W/O Cc11 .420.68100 Complex presentations -LRB- complexity = 1 -RRB- 164Appendectomy W Complicated Principal Diag W Cc2 .969.2847.25165 Appendectomy W Complicated Principal Diag W/O Cc3 .393.0454.43354 Uterine , Adnexa Proc For Non-Ovarian/Adnexal Malig W Cc2 .248.055.36355 Uterine , Adnexa Proc For Non-Ovarian/Adnexal Malig W/O Cc2 .042.1019.33357 Uterine & Adnexa Proc For Ovarian Or Adnexal Malignancy1 .8910.005.00 Diag = Diagnosis ; W/O = Without ; C.D.E. = Common Duct Exploration ; Proc = Procedure ; Malig = Malignancy ; W = With ; Cc = Complication and comorbidities Data elements included NIMs , age , gender , insurance type -LRB- Medicaid , Medicare , private , other -RRB- , hospital case mix index -LRB- CMI -RRB- , primary DRG , whether or not the admission was through emergency department -LRB- ED -RRB- , and ICD-9 procedure codes . The primary ICD-9 procedure code was used to identify both procedure -LRB- cholecystectomy , appendectomy , or hysterectomy -RRB- and type of approach -LRB- open or laparoscopic -RRB- and the primary DRG was used to differentiate simple from complex presentations in an attempt to account for intrinsic infection risks and biases towards open approaches . DRGs indicating malignancy or other complex presentations were assigned a complexity value of 1 , as shown in Table 1 . Hospital CMI was used to control for differences between and clustering within hospitals . Statistical Analysis Single and multiple logistic regression analyses were performed to quantify the associations between NIM rate and procedure , approach , patient age , gender , insurance type , complexity of presentation , ED admission status , and hospital CMI . The first model pooled all three procedures and included binary variables to adjust for the influence of each procedure on the acquisition of NIMs . Then separate models for cholecystectomy , appendectomy , and hysterectomy were constructed . Finally models were constructed for procedure and approach for wound , urinary tract , bloodstream , and respiratory tract NIMs . Results Hysterectomies comprised 43.3 % of all procedures , cholecystectomies 32.7 % , and appendectomies 24.0 % . The percentage of cholecystectomies , appendectomies , and hysterectomies that were laparoscopic was 84.7 % , 65.6 % , and 39.5 % , respectively . Unsurprisingly , fewer than one-quarter of all patients were male . Approximately 19.3 % of admissions were covered by Medicare , 7 % by Medicaid , 58.8 % by private health insurance , and the remaining 14.8 % by other types of insurance . NIM rates were defined as the number of admissions with at least one NIM divided by the total number of admissions . Of the 11,662 admissions , 337 -LRB- 2.89 % -RRB- had at least one NIM -LRB- Table 2 -RRB- . Overall , NIM rates were higher for open approaches -LRB- 4.09 % -RRB- than laparoscopic ones -LRB- 2.11 % -RRB- . NIM rates were highest for cholecystectomy -LRB- 3.57 % -RRB- , followed by appendectomy -LRB- 2.60 % -RRB- , then hysterectomy -LRB- 2.53 % -RRB- . Table 2Nosocomial infection rates by approach and procedureAdmissionsAdmissions with ≥ 1 NIMRate -LRB- % -RRB- 11,6623372.89 Approach Laparoscopic70611492 .11 Open46011884 .09 Procedure Cholecystectomy38081363 .57 Appendectomy2803732 .60 Hysterectomy50511282 .53 Approach by procedure Laparoscopic Cholecystectomy3226842 .60 Appendectomy1840422 .28 Hysterectomy1995231 .15 Open Cholecystectomy582528 .93 Appendectomy963313 .21 Hysterectomy30561053 .44 There were 399 NIMs identified in 337 admissions . Of all NIMs identified , 118 -LRB- 30 % -RRB- were from surgical wounds , 122 -LRB- 31 % -RRB- were from the urinary tract , 37 -LRB- 9 % -RRB- were from the blood , 29 -LRB- 7 % -RRB- were from the respiratory tract , and 93 -LRB- 23 % -RRB- were from other sources . At least one post-discharge NIM was identified in 136 admissions , accounting for 40 % of all admissions with a NIM . Of the 147 post-discharge NIMs , 39 % were from surgical wounds , 31 % were from the urinary tract , 7 % were from blood , and 22 % were from other sources . Of the 136 total admissions with at least one post-discharge NIM , 92 patients had NIM-associated readmissions . Univariate Analyses Simple logistic regressions examined associations between NIM rates and surgical approaches -LRB- laparoscopic or open -RRB- , and the following covariates : gender -LRB- male , female -RRB- , age -LRB- < 18 y , 18 -- 34 y , 35 -- 49 y , 50 -- 64 y , 65 -- 74 y , ≥ 75 y -RRB- , surgical procedure -LRB- cholecystectomy , appendectomy , hysterectomy -RRB- , type of insurance -LRB- private , Medicare , Medicaid , other -RRB- , complexity of admission on presentation -LRB- 0/1 -RRB- , admitted through ED -LRB- 0/1 -RRB- , and hospital CMI . The results , summarized in Table 3 , show significantly higher NIM rates for males , adults aged ≥ 65 years , patients undergoing cholecystectomy , complex admissions , and admissions covered by Medicare ; and significantly lower NIM rates for laparoscopic surgery , females , adults 18 -- 49 years old , patients undergoing hysterectomy , and those covered by private insurance . NIM rates were positively correlated with hospital CMI , but were unaffected by ED admission status . Table 3Univariate analyses of factors associated with NIMVariable CategoryNIM rate -LRB- % -RRB- OR95 % CIGenderMale3 .841.501.19 -- 1.90 Female2 .59 Age < 18 years2 .410.820.49 -- 1.3918 -- 34 years1 .500.460.32 -- 0.6535 -- 49 years2 .200.660.52 -- 0.8450 -- 64 years3 .411.250.97 -- 1.6165 -- 74 years3 .891.411.02 -- 1.95 ≥ 75 years7 .102.962.25 -- 3.90 InsurancePrivate2 .110.510.42 -- 0.65 Medicare5 .242.321.85 -- 2.92 Medicaid3 .191.120.74 -- 1.68 Other2 .770.950.70 -- 1.29 ApproachLaparoscopic2 .110.320.21 -- 0.52 Open4 .09 ProcedureCholecystectomy3 .571.411.13 -- 1.76 Appendectomy2 .600.870.67 -- 1.13 Hysterectomy2 .530.790.64 -- 0.97 CMI2 .281.59 -- 3.27 ComplexityComplex6 .312.742.14 -- 3.50 Not complex2 .40 Emergency department admissionEmergent3 .031.060.72 -- 1.55 Nonemergent2 .88 CI : confidence interval ; NIM : nosocomial infection marker ; OR : odds ratio Multivariable analyses Since ED admission status was insignificant in the univariate analysis , it was excluded from the multivariable analyses . Pairwise correlations of all remaining covariates were performed , and all pairs were reasonably uncorrelated . Therefore , all were included in the multivariable analyses . Results of multiple logistic regression models , which controlled for gender -LRB- male , female -RRB- , age -LRB- < 18 yr , 18 -- 34 yr , 35 -- 49 yr , 50 -- 64 yr , 65 -- 74 yr , ≥ 75 yr -RRB- , type of insurance -LRB- private , Medicare , Medicaid , other -RRB- , complexity of admission on presentation -LRB- 0/1 -RRB- , and hospital CMI , show that laparoscopic procedures reduced the odds of acquiring a nosocomial infections by half , but that the effect is entirely attributable to reduced infection risks in laparoscopic cholecystectomy and hysterectomy with odds ratios -LRB- ORs -RRB- of 0.34 -LRB- p < 0.01 -RRB- and 0.48 -LRB- p < 0.01 -RRB- , respectively . No change in nosocomial infection risk was found for laparoscopic appendectomy . Estimates for individual procedures are shown in Table 4 . Table 4Multivariable logistic regression analyses of factors associated with NIMVariableOdds ratio for NIMPooled -LRB- n = 11,662 -RRB- Cholecystectomy -LRB- n = 3,808 -RRB- Appendectomy -LRB- n = 2,803 -RRB- Hysterectomy -LRB- n = 5,051 -RRB- Laparoscopy0 .48 ** 0.34 ** 0.970.48 ** Type of procedure Cholecystectomy1 .87 ** -- -- -- Hysterectomy1 .05 -- -- -- Age < 18 years0 .83 -- 0.90 -- 18 -- 34 years0 .64 * 0.470.840.72 50 -- 64 years1 .222.13 * 1.000.96 65 -- 74 years1 .02 * 2.21 * 0.580.51 ≥ 75 years1 .92 ** 4.04 ** 3.310.61 Male1 .4 * 1.111.89 * -- Type of insurance Medicare1 .421.331.122.09 * Medicaid1 .453.47 ** 1.110.79 Others1 .291.531.221.32 CMI1 .63 * 1.091.312.88 * Complexity2 .45 ** NS3 .952.54 **** Statistically significant at the 1 % level * Statistically significant at the 5 % level Consistent with results of the univariate analyses , multivariable regression showed a significantly higher risk of NIMs for males , patients aged ≥ 65 years , and complexity of presentation at the time of admission . CMI was found to be significant in both univariate and multivariable analyses -LSB- OR = 1.63 -LRB- CI95 1.11 -- 2.40 -RRB- , p < 0.05 -RSB- , and this risk was significantly higher for hysterectomy patients , for whom the odds of acquiring a NIM were as high as 2.88 times that of not acquiring NIM . Analysis of the same dataset by source of infection -LRB- urinary tract , wounds , respiratory tract , bloodstream , and others -RRB- revealed that the overall infection rates at each of these sites were all statistically significantly lower for laparoscopic approaches , as summarized in Table 5 . The odds of acquiring a site-specific infection were statistically significantly lower for all sites in laparoscopic cholecystectomy and for wound sites in hysterectomy . Table 5Odds ratios by sourceUrinary tractWound Respiratory tractBloodstreamOthersOverall OR -LRB- 95 % CI -RRB- 0.61 -LRB- 0.38 -- 0.96 -RRB- 0.41 -LRB- 0.27 -- 0.62 -RRB- 0.20 -LRB- 0.08 -- 0.49 -RRB- 0.31 -LRB- 0.14 -- 0.65 -RRB- 0.52 -LRB- 0.33 -- 0.82 -RRB- By procedure Cholecystectomy0 .48 -LRB- 0.24 -- 0.97 -RRB- 0.20 -LRB- 0.11 -- 0.39 -RRB- 0.17 -LRB- 0.06 -- 0.45 -RRB- 0.23 -LRB- 0.10 -- 0.55 -RRB- 0.34 -LRB- 0.18 -- 0.64 -RRB- Appendectomy0 .83 -LRB- NS -RRB- 1.06 -LRB- NS -RRB- 0.27 -LRB- NS -RRB- Too few NIMs0 .91 -LRB- NS -RRB- Hysterectomy0 .76 -LRB- NS -RRB- 0.27 -LRB- 0.09 -- 0.79 -RRB- Too few NIMs 0.48 -LRB- NS -RRB- 0.62 -LRB- NS -RRB- OR : odds ratio , CI : confidence interval , NS : not significant There were no significant differences between laparoscopic versus open appendectomy for all sources of nosocomial infections -LRB- urine , blood , wound , respiratory , and other -RRB- . However , laparoscopic appendectomy is associated with a statistically significantly higher risk of abscess -LRB- p < 0.05 -RRB- , a finding consistent with the literature -LSB- 12 -RSB- . Forty-one percent of patients with a nosocomial infection had at least one post-discharge nosocomial infection , and 58 of 115 -LRB- 50 % -RRB- of surgical wound infections were post discharge . There were 118 readmissions associated with at least one post-discharge NIM , and post-discharge NIMs associated with readmission were significantly lower for laparoscopic approaches -LRB- p < 0.001 -RRB- . Excluding appendectomy , the odds ratio for laparoscopic versus open NIM-associated readmission was 0.346 -LRB- CI95 0.19 -- 0.63 -RRB- . Discussion This study demonstrates that laparoscopic cholecystectomy and hysterectomy reduced the overall odds of acquiring nosocomial infections from all sources by more than 50 % and reduced the odds of readmission with nosocomial infection by two-thirds . Laparoscopic appendectomy showed no differences in overall nosocomial infection risks compared to open surgery . The findings for wound infections are consistent with results from randomized trials , which have reported statistically significantly lower surgical site infection rates for laparoscopic approaches -LSB- 10 -- 12 , 20 -RSB- . This study also demonstrates statistically significant differences in source-based infection risks by procedure and approach . Specifically , wound , bloodstream , respiratory tract , urinary tract , and other nosocomial infections were all statistically significantly less likely to occur in association with laparoscopic cholecystectomy . Risks of wound infections in laparoscopic hysterectomy were also significantly lower than in open procedures . However , no differences in infection risks were found between laparoscopic and open appendectomy . It was important to stratify admission by complexity of presentation to avoid biases associated with complex presentations , higher intrinsic infection risks , and surgical approaches ; for example , primary DRGs indicating malignancy or complex presentations -LRB- Table 1 -RRB- were associated with open surgical approaches in 69 % of admissions , whereas simple presentations were associated with open approaches in only 35 % of admissions . Complex presentations are also commonly believed to be at higher risk of infections , an association that is also demonstrated in this analysis . Therefore , by controlling for presentation complexity , this analysis accounts for some of the intrinsic risk of infection as well as a bias towards open surgical approaches . The difference in patient severity between hospitals was accounted for by using CMI , and indeed CMI is significant in the univariate and multivariable models , with CMI contributing to nosocomial infection risks . Interestingly , admission through the emergency department was not significant in determining differences in nosocomial infection risks . One possible explanation is that emergency department use is a crude measure of patient severity because it may also be related to other factors such as time of day of admission and socioeconomic status . However , other variables associated with comorbidity , like age and certain payer types , were significant . Limitations While a variety of confounders were controlled for , this study is limited by the absence of certain data ; for example , antibiotic use , anesthesia scores , wound class , body mass index , prior hospitalization , and certain comorbidities -LRB- i.e. , cardiovascular status , diabetes mellitus , and immunodeficiency -RRB- were unavailable . These variables could explain additional NIM risk . Although omitted-variable bias is often a concern in multivariable modeling using retrospective databases , the similarity of findings in our univariate and multivariable analyses suggest that these results are robust . This study documents for the first time that laparoscopic hysterectomy and cholecystectomy are associated with statistically significantly lower overall risks of nosocomial infections -LRB- p < 0.01 -RRB- . Laparoscopic hysterectomy and cholecystectomy were also associated with statistically significantly fewer readmissions with nosocomial infections -LRB- p < 0.01 -RRB- . Differences in infection risks between laparoscopic and open appendectomy were not found to be statistically significant , suggesting that these differences are likely small or nonexistent . Where differences in risks have been demonstrated , future studies should be performed to quantify their effects on health care costs and length of stay . Other important directions for future research include controlling for potentially important confounders to test the robustness of our results and extending the analysis to examine the effect of laparoscopic versus open surgery on the risk of nosocomial infections for additional surgical procedures . Another interesting avenue for future research would be to examine the effect of hospital volume on the rate of nosocomial infections .

O O O B B O B O O B O B O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O B O O O O B O O O O O O O O O B O B O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O B O O O O O O O O O O O O O B B O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O B O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O B B O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O B B O O B O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O B O O O B O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O B O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O B O O O O B O O O O B O B O O B O O O O B O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O B O O O O O O O O O O B O O O O O O B O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O B O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O B O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O B O O O O O B O O O O O B O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O B B O O O O O B B O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O B O O B O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O B O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O B O O O O O O O B O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O B O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O B O O O O O O O O O O O O O O O O B B O B O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O B B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O B O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O B B O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O B O O O O B O O O O O O O O O O O O O O O O B B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O B O O O O O B O O O O O O O O O O O O O O O O O O B B O B O O O O O O B O O O O O O O O O O O O O O O O B O O O O B B O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O B B O O O O O O B B O O O O O O O O O O O O O O O O O O O B O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B B O B O O O O O O O O O B O O O O O O O B B O B O O O O O O O O O B O O O O O O O O O O O O B O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O B O O

nosocomial laparoscopic cholecystectomy appendectomy hysterectomy