bluuebunny/crossref_metadata_2025 huggingface dataset:
Optimization and Evaluation of a PCR Assay for Detecting Toxoplasmic Encephalitis in Patients with AIDS
Authors: Joseph Priya , Calderón Maritza M. , Gilman Robert H. , Quispe Monica L. , Cok Jaime , Ticona Eduardo , Chavez Victor , Jimenez Juan A. , Chang Maria C. , Lopez Martín J. , Evans Carlton A.
Published: November 2002

Toxoplasma gondii
is a common life-threatening opportunistic infection. We used experimental murine
T. gondii
infection to optimize the PCR for diagnostic use, define its sensitivity, and characterize the time course and tissue distribution of experimental toxoplasmosis. PCR conditions were adjusted until the assay reliably detected quantities of DNA derived from less than a single parasite. Forty-two mice were inoculated intraperitoneally with
T. gondii
tachyzoites and sacrificed from 6 to 72 h later. Examination of tissues with PCR and histology revealed progression of infection from blood to lung, heart, liver, and brain, with PCR consistently detecting parasites earlier than microscopy and with no false-positive results. We then evaluated the diagnostic value of this PCR assay in human patients. We studied cerebrospinal fluid and serum samples from 12 patients with AIDS and confirmed toxoplasmic encephalitis (defined as positive mouse inoculation and/or all of the Centers for Disease Control clinical diagnostic criteria), 12 human immunodeficiency virus-infected patients with suspected cerebral toxoplasmosis who had neither CDC diagnostic criteria nor positive mouse inoculation, 26 human immunodeficiency virus-infected patients with other opportunistic infections and no signs of cerebral toxoplasmosis, and 18 immunocompetent patients with neurocysticercosis. Eleven of the 12 patients with confirmed toxoplasmosis had positive PCR results in either blood or cerebrospinal fluid samples (6 of 9 blood samples and 8 of 12 cerebrospinal fluid samples). All samples from control patients were negative. This study demonstrates the high sensitivity, specificity, and clinical utility of PCR in the diagnosis of toxoplasmic encephalitis in a resource-poor setting.
Calibrated Real-Time PCR Assay for Quantitation of Human Herpesvirus 8 DNA in Biological Fluids
Authors: Broccolo Francesco , Locatelli Giuseppe , Sarmati Loredana , Piergiovanni Sara , Veglia Fabrizio , Andreoni Massimo , Buttò Stefano , Ensoli Barbara , Lusso Paolo , Malnati Mauro S.
Published: November 2002

Accurate laboratory tests for the diagnosis of active human herpesvirus 8 (HHV-8) infection are becoming essential to study the pathogenesis of HHV-8-associated tumors and for the clinical management of HHV-8-infected individuals. We have developed a highly sensitive, calibrated quantitative real-time PCR assay for the measurement of cell-free HHV-8 DNA in body fluids, based on the addition of a synthetic DNA calibrator prior to DNA extraction. The calibrator controls each sample for the presence of PCR inhibitors, determines a cutoff value of sensitivity for negative samples, and normalizes positive samples for the efficiency of DNA recovery. The assay shows a wide dynamic range of detection (between 1 and 10
6
viral genome equivalents/reaction) and a high degree of accuracy even in the presence of high amounts (up to 1 μg) of human genomic DNA. Moreover, the assay has a very high sensitivity (lower detection limit, 10 genome equivalents/ml) and a high degree of reproducibility and repeatability with a coefficient of variation (CV) of <15 and 23%, respectively. Furthermore, the use of the calibrator improves the accuracy of quantitation and decreases the intersample variability (CV, 9 and 6%, respectively). The sensitivity and specificity of the assay were tested with a series of clinical specimens obtained from patients affected by various HHV-8-related diseases, as well as from a wide number of controls. In conclusion, our calibrated real-time PCR assay provides a reliable high-throughput method for quantitation of HHV-8 DNA in clinical and laboratory specimens.
Characterization of Tn
1546
in Vancomycin-Resistant
Enterococcus faecium
Isolated from Canine Urinary Tract Infections: Evidence of Gene Exchange between Human and Animal Enterococci
Authors: Simjee S. , White D. G. , McDermott P. F. , Wagner D. D. , Zervos M. J. , Donabedian S. M. , English L. L. , Hayes J. R. , Walker R. D.
Published: November 2002

Thirty-five enterococcal isolates were recovered from dogs diagnosed with urinary tract infections at the Michigan State University Veterinary Teaching Hospital over a 2-year period (1996 to 1998). Isolated species included
Enterococcus faecium
(
n
= 13),
Enterococcus faecalis
(
n
= 7),
Enterococcus gallinarum
(
n
= 11), and
Enterococcus casseliflavus
(
n
= 4). Antimicrobial susceptibility testing revealed several different resistance phenotypes, with the majority of the enterococcal isolates exhibiting resistance to three or more antibiotics. One
E. faecium
isolate, CVM1869, displayed high-level resistance to vancomycin (MIC > 32 μg/ml) and gentamicin (MIC > 2,048 μg/ml). Molecular analysis of this isolate revealed the presence of Tn
1546
(
vanA
), responsible for high-level vancomycin resistance, and Tn
5281
carrying
aac6′-aph2"
, conferring high-level aminoglycoside resistance. Pulsed-field gel electrophoresis analysis revealed that CVM1869 was a canine
E. faecium
clone that had acquired Tn
1546
, perhaps from a human vancomycin-resistant
E. faecium
. Transposons Tn
5281
and Tn
1546
were located on two different conjugative plasmids. Sequence analysis revealed that in Tn
1546
, ORF1 had an 889-bp deletion and an IS
1216V
insertion at the 5′ end and an IS
1251
insertion between
vanS
and
vanH
. To date, this particular form of Tn
1546
has only been described in human clinical vancomycin-resistant enterococcus isolates unique to the United States. Additionally, this is the first report of a vancomycin-resistant
E. faecium
isolated from a companion animal in the United States.
Novel Single-Tube Agar-Based Test System for Motility Enhancement and Immunocapture of
Escherichia coli
O157:H7 by H7 Flagellar Antigen-Specific Antibodies
Authors: Murinda Shelton E. , Nguyen Lien T. , Ivey Susan J. , Almeida Raul A. , Oliver Stephen P.
Published: November 2002

This paper describes a novel single-tube agar-based technique for motility enhancement and immunoimmobilization of
Escherichia coli
O157:H7. Motility indole ornithine medium and agar (0.4%, wt/vol) media containing either nutrient broth, tryptone broth, or tryptic soy broth (TSBA) were evaluated for their abilities to enhance bacterial motility. Twenty-six
E. coli
strains, including 19 O157:H7 strains, 1 O157:H
−
strain, and 6 generic
E. coli
strains, were evaluated. Test bacteria were stab inoculated in the center of the agar column, and tubes were incubated at 37°C for 18 to 96 h. Nineteen to 24 of the 26 test strains (73.1 to 92.3%) were motile in the different media. TSBA medium performed best and was employed in subsequent studies of motility enhancement and H7 flagellar immunocapture. H7 flagellar antiserum (30 and 60 μl) mixed with TSBA was placed as a band (1 ml) in the middle of an agar column separating the top (3-ml) and bottom (3-ml) agar layers. The top agar layer was inoculated with the test bacterial strains. The tubes were incubated at 37°C for 12 to 18 h and for 18 to 96 h. The specificity and sensitivity of the H7 flagellar immunocapture tests were 75 and 100%, respectively. The procedure described is simple and sensitive and could be adapted easily for routine use in laboratories that do not have sophisticated equipment and resources for confirming the presence of H7 flagellar antigens. Accurate and rapid identification of H7 flagellar antigen is critical for the complete characterization of
E. coli
O157:H7, owing to the immense clinical, public health, and economic significance of this food-borne pathogen.
Limited Diversity among Human Isolates of
Bartonella henselae
Authors: Dillon B. , Valenzuela J. , Don R. , Blanckenberg D. , Wigney D. I. , Malik R. , Morris A. J. , Robson J. M. , Iredell J.
Published: November 2002

A study of 59 isolates of
Bartonella henselae
reveals relatively limited diversity among those of human origin (
n
= 28). Either of two distinct alleles of both
gltA
and 16S ribosomal DNA (rDNA) was found in all isolates, with a high level of congruity between 16S and
gltA
inheritance among proven human pathogens. Human isolates from all over Eastern Australia were most commonly 16S rDNA (Bergmans) type I, with the same
gltA
allele as the type strain (Houston-1). Comparable feline isolates were more commonly 16S type II, with less congruity of inheritance between 16S and
gltA
alleles. Previously described arbitrarily primed PCR and
Eag
I-
Hha
I infrequent restriction site PCR fingerprinting techniques separated
Bartonella
species effectively but lacked discriminating power within
B. henselae.
Examination of the 16-23S intergenic spacer region revealed for several strains several point mutations as well as a repeat sequence of unknown significance which is readily detected by
Hae
III restriction fragment length polymorphism analysis. The bacteriophage-associated
papA
gene was present in all isolates. Enterobacterial repetitive intergenic consensus PCR proved to be a useful and robust typing tool and clearly separated human isolates (including imported strains) from the majority of feline isolates. Our data are consistent with published evidence and with previous suggestions of intragenomic rearrangements in the type strain and suggest that human isolates come from a limited subset of
B. henselae
strains. They strengthen arguments for careful exploration of genotype-phenotype relationships and for the development of a multilocus enzyme electrophoresis and multilocus sequence typing-based approach to the phylogeny of
B. henselae
.
Detection of Mycobacterial DNA in Andean Mummies
Authors: Konomi Nami , Lebwohl Eve , Mowbray Ken , Tattersall Ian , Zhang David
Published: November 2002

The identification of genetic material from pathogenic organisms in ancient tissues provides a powerful tool for the study of certain infectious diseases in historic populations. We have obtained tissue samples from the genital areas of 12 mummies in the American Museum of Natural History collection in New York, N.Y. The mummies were excavated in the Andes Mountain region of South America, and radiocarbon dating estimates that the mummies date from
a.d.
140 to 1200. DNAs were successfully extracted from all tissues and were suitable for PCR analysis. PCRs were carried out to detect
Mycobacterium tuberculosis
complex and mycobacteria other than
M. tuberculosis
(MOTB).
M. tuberculosis
complex was detected in 2 out of 12 samples, and MOTB were detected in 7 samples. This study confirmed the adequate preservation of genetic material in mummified tissues and the existence of mycobacteria, including
M. tuberculosis
, in historic populations in South America.
New Technique (the NOW Test) for Rapid Detection of
Streptococcus pneumoniae
in the Nasopharynx
Authors: Faden Howard , Heimerl Michael , Goodman Gail , Winkelstein Peter , Varma Chelikani
Published: November 2002

Although the NOW test was originally introduced as a rapid pneumococcal antigen detection test for use with urine samples, it was successfully adapted to nasopharyngeal samples in the present study. The sensitivity, specificity, positive predictive value, and negative predictive value of the test were 92.2, 97.7, 95.9, and 95.5%, respectively. These results demonstrate that nasopharyngeal colonization with
Streptococcus pneumoniae
can be documented within 15 min of sample collection.
Comparison of the Borrelia DotBlot G, MarDx, and VIDAS Enzyme Immunoassays for Detecting Immunoglobulin G Antibodies to
Borrelia burgdorferi
in Human Serum
Authors: Jespersen Deborah J. , Smith Thomas F. , Rosenblatt Jon E. , Cockerill Franklin R.
Published: November 2002

Three enzyme immunoassays, Borrelia DotBlot G (GenBio, San Diego, Calif.), MarDx EIA (MarDx Diagnostics, Inc., Carlsbad, Calif.), and VIDAS (bioMérieux, St. Louis, Mo.) were compared for their ability to detect immunoglobulin G antibodies to
Borrelia burgdorferi
in 100 human serum samples. The “gold standard” positive result for each of these samples was determined by Western immunoblot analysis (MarDx Marblot Test System) (
n
= 99) or clinical signs and symptoms suggestive of Lyme disease with other laboratory results positive for
B. burgdorferi
(
n
= 1). Based on these criteria for a gold standard positive result, 29 of the 100 samples tested were considered true positives and 71 were considered true negatives. The following sensitivities and specificities were noted, respectively, for each method: Borrelia DotBlot G, 93 and 90%; MarDx, 100 and 35%; and VIDAS, 100 and 92%. Because of high sensitivity and specificity and ease of use, the VIDAS test is an appealing method, especially for laboratories that perform high volumes of tests. The high sensitivity but low specificity of the MarDx method compared to the VIDAS and Borrelia DotBlot G methods requires that Western blot confirmatory tests be performed frequently. The Borrelia DotBlot G method has acceptable specificity but appears to lack sensitivity when compared to the VIDAS and MarDx methods.
Injuries in professional horse racing in Great Britain and the Republic of Ireland during 1992–2000
Authors: Turner M , McCrory P , Halley W
Published: November 2002

Professional horse racing is an exciting and demanding sport with high injury rates. Surprisingly few epidemiological studies have been published and no prospective studies have been reported. This paper reviews the literature and provides a detailed breakdown of injuries in Great Britain and the Republic of Ireland for 1992–2000. The introduction of protective equipment is discussed and evidence for its effectiveness put forward.
Ophthalmological findings in elite amateur Turkish boxers
Authors: Hazar M , Beyleroglu M , Subasi M , Or M
Published: November 2002

Objective:To evaluate the nature and incidence of pathological ocular conditions in boxing.

Methods:A group of 20 active, elite, amateur, asymptomatic Turkish boxers were examined and compared with a control group composed of 20 age matched sportsmen who were not boxers. The boxers had been actively boxing for 5–20 years (mean 9.65), were aged 16–34 (mean 22.25), and weighed 51–91 kg (mean 73.07). They had been involved in 67–500 fights (mean 143.8), with 5–40 losses (mean 17.75). All were championship title holders at the national, European, Olympic, or World level.

Results:An atrophic retinal hole was found in one boxer, which was treated with laser prophylaxis. In this series, the incidence of traumatic eye injuries was much lower than in the literature. Possible reasons are their young age, their division being mainly lightweight or middleweight, few fight losses, being elite boxers, their fighting distance being near or distant, protective methods, and racial factors.

Conclusion:Boxing does not appear to be as hazardous to the eyes as previously reported, but prospective, longitudinal studies need to be carried out, including boxers of all divisions, ages, boxing experience, and level, and using various methods of fighting and protection. It should be made mandatory for all boxers to have a complete ophthalmological examination at the beginning of their careers and periodically thereafter.
The Football Association Medical Research Programme: an audit of injuries in professional football—analysis of preseason injuries
Authors: Woods C , Hawkins R , Hulse M , Hodson A
Published: November 2002

Objectives:To conduct a detailed analysis of preseason football injuries sustained in English professional football over two competitive seasons.

Methods:Club medical staff at 91 professional football clubs annotated player injuries. A specific injury audit questionnaire was used together with a weekly form that documented each club’s current injury status.

Results:17% (1025) of the total number of injuries over the two seasons were sustained during the preseason, the mean number of days absent per injury was 22.3 days. Younger age groups (17–25 yrs) were more likely to sustain a preseason injury than more experienced players (26–35+) (p<0.01). There were relatively more “slight” and “minor” injuries (as defined in the methodology), overuse, and tendon related injuries sustained during preseason compared to the in season (p<0.01). The thigh (23%), knee (17%), and ankle (17%) were the most common locations for injuries during the preseason, there was a relatively greater number of lower leg injuries (15%) during the preseason (p<0.05). Achilles tendonitis was most prevalent in the preseason, with 33% of all Achilles related injuries sustained during this period (p<0.01). Muscle strains were the most common injury during preseason (37%). Rectus femoris muscle strains were observed twice as frequently during the preseason relative to the in season (p<0.01). Ligament sprains were the second most common injury during preseason (19%). Non-contact mechanisms were the cause of significantly more injuries during the preseason (p<0.01), with relatively more preseason injuries sustained while running or shooting (p<0.01). For 70% of the injuries reported during the preseason, the ground condition was described as dry.

Conclusions:Players are at a greater risk of slight and minor injuries, overuse injuries, lower leg injuries (especially the Achilles tendon) and rectus femoris strains during the preseason period. Prevention of preseason injury is important to ensure availability of players for the commencement of the season and to decrease the risk of injury later in the season, we recommend the implementation of a risk management policy for this purpose. Areas requiring further investigation include methods of prevention for the common preseason injuries that have been identified, a detailed analysis of preseason and closed season training programmes, and a smaller study involving exposure data.
Complete rupture of the triceps brachii muscle
Authors: Singh R K , Pooley J
Published: November 2002

The case is reported of a professional ice hockey player who sustained an intramuscular rupture of all three heads of the triceps brachii muscle. After surgical repair, he made a complete recovery and was able to return to professional ice hockey.
Groin pain in sportsmen is not always musculotendinous in origin
Authors: Gomez K F , Dawson R , Davies S G , Foster M E
Published: November 2002

The case is reported of a young rugby player who presented with pubic tenderness not caused by any muscular injury or direct trauma but by a retropubic abscess.
Late complications following treatment for severe aplastic anemia (SAA) with high-dose cyclophosphamide (Cy): follow-up of a randomized trial
Authors: Tisdale John F. , Maciejewski Jaroslaw P. , Nuñez Olga , Rosenfeld Stephen J. , Young Neal S.
Published: November 2002

High-dose cyclophosphamide (Cy) has been promoted as curative therapy for severe aplastic anemia (SAA). However, our randomized trial comparing antithymocyte globulin (ATG) and Cy was terminated early because of excess morbidity/early mortality in the Cy arm. We now report analysis of secondary endpoints at a median of 38 months. Relapse occurred in 6 (46%) of 13 responders in the ATG arm versus 2 (25%) of 8 in the Cy arm (P = .38). Five (31%) of 16 patients in the ATG arm and 4 (27%) of 15 patients in the Cy arm had evidence of paroxysmal nocturnal hemoglobinuria (PNH) at diagnosis, with no substantial change in the overall percentage of glycophosphatidyl inositol (GPI)–anchored protein-deficient neutrophils over extended follow-up in individual patients in either arm. Bone marrow cytogenetic abnormalities have been observed among surviving patients in both arms (2 of 14 ATG versus 1 of 12 Cy,P = .70). High-dose Cy does not prevent relapse or clonal evolution in SAA.
Dysregulated lymphocyte proliferation and differentiation in patients with rheumatoid arthritis
Authors: Ponchel Frederique , Morgan Ann W. , Bingham Sarah J. , Quinn Mark , Buch Maya , Verburg Robert J. , Henwood Judy , Douglas Susan H. , Masurel Aurelie , Conaghan Philip , Gesinde Moji , Taylor Julia , Markham Alexander F. , Emery Paul , van Laar Jacob M. , Isaacs John D.
Published: November 2002

Rheumatoid arthritis (RA) is a chronic, inflammatory disease of the synovium of uncertain pathogenesis. A number of phenotypic and functional T-cell defects have been described in RA, including abnormal clonal expansions and suppressed proliferative responses, which suggest a defect in T-cell differentiation. Here, we show that RA patients possess fewer naive CD4+ T cells than healthy controls. Furthermore, a smaller proportion of these cells contains a T-cell receptor excision circle (TREC). Patients with RA also have unusual populations of T cells. These include immature cells characterized as CD45RBbrightCD45RA+CD62L− by flow cytometry and a large population that coexpresses CD45RA and CD45RO. These cells are hyperresponsive to mitogen and TCR stimulation when compared to naive cells. Additionally, an unusual putative central memory subset expressing CD62L, but not CD45RA, appears in RA patients at the expense of more typical cells. Levels of C-reactive protein correlate inversely with the TREC content of naive T cells and positively with the sizes of naive and immature atypical T-cell subsets. These data suggest that inflammation drives proliferation of naive T cells in RA and encourages their differentiation into atypical, hyperresponsive progeny. TREC content of individual naive and atypical T-cell subsets suggests an ontogeny consistent with this hypothesis. These studies provide further evidence of a T-cell differentiation defect in RA, which could explain some of the well-characterized immunologic features of the disease.
Distal elements are critical for human CD34 expression in vivo
Authors: Okuno Yutaka , Huettner Claudia S. , Radomska Hanna S. , Petkova Victoria , Iwasaki Hiromi , Akashi Koichi , Tenen Daniel G.
Published: November 2002

The elements regulating gene expression in hematopoietic stem cells are still poorly understood. We previously reported that a 141-kilobase (kb) human CD34 transgene confers properly regulated human CD34 expression in transgenic mice. A construct with only the human CD34 promoter and 3′ enhancer region is not sufficient, suggesting that critical distal elements are necessary for expression of the human CD34 gene. To further localize such elements, we analyzed deletion constructs of the human CD34 gene and evaluated their function in transgenic mice. Constructs harboring as little as 18 kb of 5′ and 26 kb of 3′ human CD34 flanking sequence conferred human expression in tissues of transgenic mice with a pattern similar to that of the 141-kb human transgene. In contrast, a construct harboring 10 kb of 5′ and 17 kb of 3′ human CD34 flanking sequence gave no expression. These data demonstrate that regions between 10 to 18 kb upstream and/or 17 to 26 kb downstream of the human CD34 gene contain critical elements for human CD34 expression in vivo. Further functional analysis of these regions in transgenic mice will be crucial for understanding CD34 gene expression in hematopoietic stem and progenitor cells.
CD45 tyrosine phosphatase inhibits erythroid differentiation of umbilical cord blood CD34+ cells associated with selective inactivation of Lyn
Authors: Harashima Akira , Suzuki Motoyuki , Okochi Ayumi , Yamamoto Mayuko , Matsuo Yoshinobu , Motoda Ryuichi , Yoshioka Tamotsu , Orita Kunzo
Published: November 2002

CD45 is a membrane-associated tyrosine phosphatase that dephosphorylates Src family kinases and Janus kinases (JAKs). To clarify the role of CD45 in hematopoietic differentiation, we examined the effects of anti-CD45 monoclonal antibody NU-LPAN on the proliferation and differentiation of umbilical cord blood CD34+ cells. NU-LPAN showed a prominent inhibition of the proliferation of CD34+ cells induced by the mouse bone marrow stromal cell line MS-5 or erythropoietin (EPO). However, NU-LPAN did not affect the proliferation induced by interleukin 3. NU-LPAN also inhibited MS-5–induced or EPO-induced erythroid differentiation of CD34+ cells. The cells stimulated with EPO in the presence of NU-LPANmorphologically showed differentiation arrest at the stage of basophilic erythroblasts after 11 days of culture, whereas the cells treated with EPO without NU-LPAN differentiated into mature red blood cells. The Src family kinase Lyn and JAK2 were phosphorylated when erythroblasts obtained after 4 days of culture of CD34+ cells in the presence of EPO were restimulated with EPO. Overnight NU-LPAN treatment before addition of EPO reduced the phosphorylation of Lyn but not that of JAK2. Simultaneously, the enhancement of Lyn kinase activity after restimulation with EPO was reduced by NU-LPAN treatment. These results indicate selective inactivation of Lyn by CD45 activated with NU-LPAN and could partly explain the inhibitory mechanism on erythropoiesis exhibited by EPO. These findings suggest that CD45 may play a pivotal role in erythropoiesis.
Human myeloma cells stimulate the receptor activator of nuclear factor-κB ligand (RANKL) in T lymphocytes: a potential role in multiple myeloma bone disease
Authors: Giuliani Nicola , Colla Simona , Sala Roberto , Moroni Matteo , Lazzaretti Mirca , La Monica Silvia , Bonomini Sabrina , Hojden Magda , Sammarelli Gabriella , Barillè Sophie , Bataille Regis , Rizzoli Vittorio
Published: November 2002

The biologic mechanisms involved in the pathogenesis of multiple myeloma (MM) bone disease are not completely understood. Recent evidence suggests that T cells may regulate bone resorption through the cross-talk between the critical osteoclastogenetic factor, receptor activator of nuclear factor-κB ligand (RANKL), and interferon γ (IFN-γ) that strongly suppresses osteoclastogenesis. Using a coculture transwell system we found that human myeloma cell lines (HMCLs) increased the expression and secretion of RANKL in activated T lymphocytes and similarly purified MM cells stimulated RANKL production in autologous T lymphocytes. In addition, either anti–interleukin 6 (anti–IL-6) or anti–IL-7 antibody inhibited HMCL-induced RANKL overexpression. Consistently, we demonstrated that HMCLs and fresh MM cells express IL-7 mRNA and secrete IL-7 in the presence of IL-6 and that bone marrow (BM) IL-7 levels were significantly higher in patients with MM. Moreover, we found that the release of IFN-γ by T lymphocytes was reduced in presence of both HMCLs and purified MM cells. Furthermore, in a stromal cell–free system, osteoclastogenesis was stimulated by conditioned medium of T cells cocultured with HMCLs and inhibited by recombinant human osteoprotegerin (OPG; 100 ng/mL to 1 μg/mL). Finally, RANKL mRNA was up-regulated in BM T lymphocytes of MM patients with severe osteolytic lesions, suggesting that T cells could be involved at least in part in MM-induced osteolysis through the RANKL overexpression.
In vivo administration of vascular endothelial growth factor (VEGF) and its antagonist, soluble neuropilin-1, predicts a role of VEGF in the progression of acute myeloid leukemia in vivo
Authors: Schuch Gunter , Machluf Marcelle , Bartsch Georg , Nomi Masashi , Richard Henri , Atala Anthony , Soker Shay
Published: November 2002

Recent findings implied that the progression of hematologic malignancies, like that of solid tumors, is dependent on neovascularization. Recent studies on patients with acute myeloid leukemia (AML) showed increased levels of leukocyte-associated vascular endothelial growth factor (VEGF) and neovascularization of the bone marrow. Murine (32D, M1) and human (HEL, U937, and UKE-1) leukemic cell lines and freshly isolated leukemic cells were analyzed for the expression of VEGF and VEGF receptor mRNA. The expression of VEGF and VEGF receptors KDR and neuropilin-1 (NRP-1) was detected in these cells. In a murine chloroma model, delivery of VEGF165using microencapsulation technology resulted in enhanced tumor growth and vascularization, whereas treatment with a VEGF antagonist soluble NRP-1 (sNRP-1) inhibited tumor angiogenesis and growth. In a systemic leukemia model, survival of mice injected with adenovirus (Ad) encoding for Fc-sNRP-1 (sNRP-1 dimer) was significantly prolonged as compared with mice injected with Ad-LacZ. Further analyses showed a reduction in circulating leukemic cells and infiltration of liver and spleen as well as bone marrow neovascularization and cellularity. Taken together, these results demonstrate that angiogenic factors such as VEGF promote AML progression in vivo. The use of VEGF antagonists as an antiangiogenesis approach offers a potential treatment for AML. Finally, our novel in vivo drug delivery model may be useful for testing the activities of other peptide antiangiogenic factors.
Early Detection of Toxicity and Adjustment of Ongoing Clinical Trials: The History and Performance of the North Central Cancer Treatment Group’s Real-Time Toxicity Monitoring Program
Authors: Goldberg Richard M. , Sargent Daniel J. , Morton Roscoe F. , Mahoney Michelle R. , Krook James E. , O’Connell Michael J.
Published: November 2002

ABSTRACT: Prospective clinical trials are the gold standard for evidence-based methodology used to support changes in the practice of medicine. Clinical researchers, regulatory agencies, payers, and the public embrace the conduct of phase I, II, and III clinical trials as integral to improving patient care. The National Cancer Institute (NCI) funds a number of cooperative oncology groups to conduct such clinical trials in the United States. In order to protect enrolling patients, the NCI requires expedited reporting to allow rapid identification of severe side effects on NCI-sponsored clinical trials. However, chemotherapy drugs frequently cause predictable side effects, the rapid reporting of which would potentially overwhelm the system. This article describes the development and documents the performance of a real-time toxicity reporting system implemented by the North Central Cancer Treatment Group. The goal of this system is to supplement the currently required NCI adverse event monitoring procedures and to permit study teams to identify the need to modify ongoing clinical trials. The system has proven its value in the monitoring of phase II and III trials, including trial N9741, a three-arm, phase III, advanced colorectal cancer chemotherapy study exploring combinations of irinotecan, oxaliplatin, and fluorouracil. We believe the methods described present opportunities for improving patient safety in clinical research.
Long-Term Enalapril Therapy for Left Ventricular Dysfunction in Doxorubicin-Treated Survivors of Childhood Cancer
Authors: Lipshultz Steven E. , Lipsitz Stuart R. , Sallan Stephen E. , Simbre Valeriano C. , Shaikh Seema L. , Mone Suzanne M. , Gelber Richard D. , Colan Steven D.
Published: November 2002

PURPOSE: A common late effect of doxorubicin therapy for childhood cancer is reduced left-ventricular (LV) wall thickness resulting in elevated LV afterload and depressed LV function. Many children are given angiotensin-converting enzyme inhibitors, which have been studied primarily in adults. We document the long-term effects of angiotensin-converting enzyme inhibitors in doxorubicin-treated survivors of childhood cancer.

PATIENTS AND METHODS: In this retrospective study, we reviewed records of 18 children who had regular echocardiographic examinations during enalapril therapy (mean age at cancer diagnosis, 8 years; mean time between completion of doxorubicin therapy and start of enalapril, 7 years; median follow-up since the start of enalapril, 10 years).

RESULTS: Over the first 6 years of enalapril therapy, there was progressive improvement toward normal values in LV dimension, afterload, fractional shortening, and mass, but all these parameters deteriorated between 6 and 10 years. LV wall thickness deteriorated throughout the study period, as did LV contractility and systolic blood pressure. Diastolic blood pressure fell slightly. By 6 years on enalapril, all six patients who had had congestive heart failure at the start of enalapril therapy had either died or undergone cardiac transplantation, compared with three of the 12 asymptomatic patients.

CONCLUSION: In doxorubicin-treated long-term survivors of childhood cancer, enalapril-induced improvement in LV structure and function is transient. The primary defect, which is LV wall thinning, continues to deteriorate, and thus the short-term improvement was mostly related to lowered diastolic blood pressure.
Kinetics of lactose hydrolysis by β‐galactosidase of Kluyveromyces lactis immobilized on cotton fabric
Authors: Zhou Quinn Zhengkun , Chen Xiao Dong , Li Xuemei
Published: November 2002

A mathematic model for describing the Michaelis‐Menten‐type reaction kinetics with product competitive inhibition and side‐reaction is proposed. A multiresponse nonlinear simulation program was employed to determine the coefficients of a four‐parameter rate expression. The rate expression was compared with the conventional Michaelis‐Menten reaction rate models with and without product inhibition. Experimental data were obtained using β‐galactosidase of Kluyveromyces lactis immobilized on cotton fabric in a batch system at a temperature of 37°C and at various initial concentrations of dissolved lactose ranging from 3–12.5% (w/v). The reaction is followed by concentration changes with time in the tank. Samples were obtained after the outlet stream of the packed bed reactor is mixed in a well‐stirred tank. High‐performance liquid chromatography (HPLC) was applied to monitor the concentrations of all the sugars (reactants as well as products). The four‐parameter rate model is featured with a term to describe the formation of trisaccharides, a side‐reaction of the enzymatic hydrolysis. The proposed model simulates the process of lactose hydrolysis and the formation of glucose and galactose, giving better accuracy compared with the previous models. © 2002 Wiley Periodicals, Inc. Biotechnol Bioeng 81: 127–133, 2003.
Ultra scale‐down approach for the prediction of full‐scale recovery of ovine polycolonal immunoglobulins used in the manufacture of snake venom‐specific Fab fragment
Authors: Neal G. , Christie J. , Keshavarz‐Moore E. , Shamlou P. Ayazi
Published: November 2002

In this article, we describe a new approach that allows the prediction of the performance of a large‐scale integrated process for the primary recovery of a therapeutic antibody from an analysis of the individual unit operations and their interactions in an ultra scale‐down mimic of the process. The recovery process consisted of four distinct unit operations. Using the new approach we defined the important engineering parameters in each operation that impacted the overall recovery process and in each case verified its effect by a combination of modelling and experimentation. Immunoglobulins were precipitated from large volumes of dilute blood plasma and the precipitated flocs were recovered by centrifugal separation from the liquor containing contaminating proteins, including albumin. The fluid mechanical forces acting on the precipitate and the time of exposure to these forces were used to define a time‐integrated fluid stress. This was used as a scaling factor to predict the properties of the precipitated flocs at large scale. In the case of centrifugation, the performance of a full‐scale disc stack centrifuge was predicted. This was achieved from a computational fluid dynamics (CFD) analysis of the flow field in the centrifuge coupled with experimental data obtained from the precipitated immunoglobulin flocs using the scale‐down precipitation tank, a rotating shear device, and a standard swing‐out rotor centrifuge operating under defined conditions. In this way, the performance of the individual unit operations, and their linkage, was successfully analysed from a combination of modelling and experiments. These experiments required only millilitre quantities of the process material. The overall performance of the large‐scale process was predicted by tracking the changes in physical and biological properties of the key components in the system, including the size distribution of the antibody precipitates and antibody activity through the individual unit operations in the ultra scale‐down process flowsheet. © 2002 Wiley Periodicals, Inc. Biotechnol Bioeng 81: 149–157, 2003.
Characterization of gas–liquid mass transfer phenomena in microtiter plates
Authors: Hermann Robert , Lehmann Mathias , Büchs Jochen
Published: November 2002

Gas–liquid mass transfer properties of shaken 96‐well microtiter plates were characterized using a recently described method. The maximum oxygen transfer capacity (OTRmax), the specific mass transfer area (a), and the mass transfer coefficient (kL) in a single well were determined at different shaking intensities (different shaking frequencies and shaking diameters at constant filling volume) and different filling volumes by means of sulfite oxidation as a chemical model system. The shape (round and square cross‐sections) and the size (up to 2 mL maximum filling volume) of a microtiter plate well were also considered as influencing parameters. To get an indication of the hydrodynamic behavior of the liquid phase in a well, images were taken during shaking and the liquid height derived as a characteristic parameter. The investigations revealed that the OTRmax is predominantly dependent on the specific mass transfer area (a) for the considered conditions in round‐shaped wells. The mass transfer coefficient (kL) in round‐shaped wells remains at a nearly constant value of about 0.2 m/h for all shaking intensities, thus within the range reported in the literature for surface‐aerated bioreactors. The OTRmax in round‐shaped wells is strongly influenced by the interfacial tension, determined by the surface tension of the medium used and the surface properties of the well material. Up to a specific shaking intensity the liquid surface in the wells remains horizontal and no liquid movement can be observed. This critical shaking intensity must be exceeded to overcome the surface tension and, thus, to increase the liquid height and enlarge the specific mass transfer area. This behavior is solely specific to microtiter plates and has not yet been observed for larger shaking bioreactors such as shaking flasks. In square‐shaped microtiter plate wells the corners act as baffles and cause a significant increase of OTRmax, a, and kL. An OTRmax of up to 0.15 mol/L/h can be reached in square‐shaped wells. © 2002 Wiley Periodicals, Inc. Biotechnol Bioeng 81: 178–186, 2003.
Immunoaffinity removal of xenoreactive antibodies using modified dialysis or microfiltration membranes
Authors: Karoor Sujatha , Molina Jeanette , Buchmann Cody R. , Colton Clark , Logan John S. , Henderson Lee W.
Published: November 2002

Hyperacute rejection following xenogeneic transplantation in primates is mediated by naturally occurring IgM antibodies, which are specifically directed to α‐Galactosyl residues on many nonprimate mammalian cells. Current approaches to remove these anti‐αGal IgM include plasmapheresis followed by immunoaffinity adsorption on bead columns using synthetic Gal epitopes, which requires two pieces of complex equipment. In this study, we explored the use of immunoaffinity adsorption with hollow fiber microporous or dialysis membranes to which a synthetic αGal trisaccharide ligand is bound. Covalent attachment of ligand directly to the surface produced negligible binding, but use of long‐chain polyamines as reactive spacers yielded binding densities for anti‐αGal IgM as high as 89 mg/mL membrane volume in breakthrough curve experiments with microporous nylon membranes having an internal surface area of 4.2 m2/mL membrane volume. A crossflow microfilter fabricated from the membranes described in this study and having about 0.4 m2 luminal surface area would be able to carry out plasma separation and immunoadsorption in a single device with a large excess of binding capacity to ensure that all plasma that filters across the device and is returned to a human patient is essentially free of anti‐αGal IgM. We conclude that immunoaffinity removal of xenoreactive antibodies using microfiltration hollow fiber membranes is feasible and has potential advantages of efficiency and simplicity for clinical application. © 2002 Wiley Periodicals, Inc. Biotechnol Bioeng 81: 134–148, 2003.
Size and shape characteristics of SDH rings
Authors: Naldi M. , Agostini L.
Published: November 2002

A number of issues concerning the SDH networks (e.g. performance analysis of traffic management policies and statistical cost evaluation) may require a simulation approach that takes into account the size and shape of SDH rings. As a statistical support for the modelling of SDH rings for these purposes, the results of an analysis conducted on a real sample network of moderate‐to‐large dimensions are reported in this paper. Some items (e.g. the no. of nodes per ring, the area enclosed by the ring, and the perimeter vs area relationship) are selected to characterize the rings' size and shape. For each item the relevant statistics are supplied. In addition, models are proposed for the most relevant characteristics. Bounds are obtained for the average internodal distance on a ring. A log‐normal probability model is suggested for the internodal distances and a mixture of two Gaussian components is obtained for the distance between the ring's nodes and the ring's centroid. A power regression curve is instead used to fit the perimeter vs area experimental data, showing that the shape of SDH rings could be well modelled by an ellipse. Copyright © 2002 John Wiley & Sons, Ltd.
A novel delineation mechanism for the ATM adaptation layer 2 over wireless ATM networks
Authors: Villasenor‐Gonzalez Luis , Orozco‐Barbosa Luis , Lamont Louise
Published: November 2002

The ATM adaptation layer 2 (AAL2) has been standardized by the ITU‐T for the support of low data rate and delay‐sensitive applications, such as voice, over ATM networks. One of the main characteristics of the AAL2 standard is the support for multiplexing information at the expense of introducing a new frame structure inside the payload of the ATM cells. The AAL2 standard introduces a mechanism for the delineation of the AAL2 packets and it has been found that the delineation mechanism reduces the performance of AAL2 in the presence of channels with high bit error rates. In this paper, a novel delineation mechanism is proposed for AAL2 to be used over highly error‐prone channels, such as wireless links. The proposed mechanism improves the performance (i.e. reduces packet loss) of the AAL2 standard in the presence of bit errors and in some cases reduces the overhead required for the delineation of the AAL2 packets. Copyright © 2002 John Wiley & Sons, Ltd.
Fine‐needle aspiration cytology of extranodal natural killer/T‐cell lymphoma
Authors: Jiménez‐Heffernan José A. , González‐Peramato Pilar , Perna Cristian , Alvarez‐Ferreira Javier , López‐Ferrer Pilar , Viguer José M.
Published: November 2002

Extranodal NK/T‐cell lymphoma, nasal type, is a predominantly extranodal lymphoma characterized histologically by prominent necrosis, angiocentric growth, and vascular destruction. Only one report describing its fine‐needle aspiration (FNA) cytologic features is available and shows highly unusual findings for a lymphoma. The present case concerns a 58‐yr‐old patient that presented with a soft tissue mass of the thigh in addition to an ulcerative lesion of the palate and nodular hepatic and splenic lesions. FNA cytology of the thigh tumor was interpreted as a malignant mesenchymal lesion (sarcoma). The subsequent pathologic study revealed an NK/T‐cell lymphoma. Our findings are very similar to those previously reported. They were highly unusual for a lymphoma and consisted of polymorphic, round to spindle neoplastic cells distributed in irregular aggregates, and single cells. No significant number of lymphoglandular bodies were present. Diagn. Cytopathol. 2002;27:371–374. © 2002 Wiley‐Liss, Inc.
Utility of fine‐needle aspiration in the diagnosis of primary osteosarcoma
Authors: Dodd Leslie G. , Scully Sean P. , Cothran R. Lee , Harrelson John M.
Published: November 2002

Fine‐needle aspiration (FNA) is a reliable, safe, and cost‐effective procedure with a well‐established role in the diagnosis of various solid tissue neoplasms. The role of FNA in the diagnosis of primary bone tumors, including osteosarcoma (OGS), is controversial and has yet to be established. We reviewed our experience with the use of FNA as a diagnostic technique over the past 8 yr at our institution. Diagnosis was conclusive in 26 (65%) of 40 patients, 18 of whom went to neoadjuvant therapy and/or resection based solely on the FNA interpretation of either “high grade sarcoma” or “osteosarcoma.” Of the remaining 14 (25%) patients, 12 had inconclusive diagnosis and two (5%) were false‐negatives. An inconclusive diagnosis was most likely to be an inadequate or paucicellular aspirate, seen in six (15%) patients. An additional six patients had variants of osteosarcoma (four chondroid, one “giant cell rich,” one parosteal) that made definitive diagnosis impossible. The two that were incorrectly classified were diagnosed as fracture callus and plasmacytoma. FNA is an accurate and cost‐effective tool for the initial diagnosis of primary osteosarcoma with a sensitivity of 65% and accuracy of 95%. Inconclusive diagnoses are likely to be due to insufficient sample cellularity or the presence of OGS variant. In our experience, FNA is sufficient to provide the diagnosis of OGS prior to definitive treatment when interpreted in conjunction with imaging studies and clinical findings. In those cases where FNA fails to yield a diagnostic sample, a traditional biopsy can be performed. Diagn. Cytopathol. 2002;27:350–353. © 2002 Wiley‐Liss, Inc.
Cytologic features of metastatic nasopharyngeal carcinoma
Authors: Mohanty Sambit K. , Dey Pranab , Ghoshal Sushmita , Nahar Saikia Uma
Published: November 2002

The aim of the study was to describe the cytologic features of metastatic nasopharyngeal carcinomas (NPC) to lymph node, which would help in arriving at a definite diagnosis, particularly in cases of occult primary with predominant finding of cervical lymphadenopathy. This is a retrospective study over a period of 2 yr (January 2000 to December 2001) and included 15 cases of NPC metastasizing to cervical lymph nodes. All cases had histopathologic diagnosis of primary NPC. Two independent observers studied detailed cytomorphology of these cases. The FNAC smears were cellular and except for three cases revealed predominantly discohesive malignant cells admixed with lymphocytes. Cellular degeneration and naked tumor cell nuclear were seen in all cases. The tumor cells showed mild to moderate nuclear pleomorphism, vesicular chromatin, and scanty cytoplasm with ill‐defined cytoplasmic borders. All 15 cases were typed according to the World Health Organization (WHO) classification as types I, II, and III NPCs. There were two cases of type I NPCs, five cases of type II, and eight cases of type III pattern. FNAC of metastatic NPC in lymph node is a valuable diagnostic modality in suggesting the site of origin of the tumor. Furthermore, it is also helpful to exclude residual disease and recurrence in a treated patient with lymphadenopathy. Diagn. Cytopathol. 2002;27:340–342. © 2002 Wiley‐Liss, Inc.