The Effects of Probiotics on Metabolic Biomarkers, Inflammation, and the Antioxidant System in Patients With Type 2 Diabetes Mellitus
Phase: NA
Conditions: Diabetes Mellitus (Type 2), Inflammation, Probiotics, Antioxidant Effect, Glucose Metabolism, Lipid Metabolism
Sponsor: Ege University

 
 
 
OFFICIAL TITLE: 
 
THE EFFECTS OF PROBIOTICS ON 
METABOLIC BIOMARKERS, 
INFLAMMATION, AND THE 
ANTIOXIDANT SYSTEM IN PATIENTS 
WITH TYPE 2 DIABETES MELLITUS 
Date: 17.04.2018 
 

Background and Object1ves 
Type 2 d(abetes mell(tus (T2DM) prevalence has (ncreased alarm(ngly due to obes(ty and 
sedentary l(festyle trends [1, 2]. T2DM causes end-stage renal fa(lure, bl(ndness, and lower l(mb 
amputat(ons through progress(ve atheroscleros(s, lead(ng to 3-fold (ncreased card(ovascular 
events and 2-fold (ncreased cerebrovascular events [3, 4]. 
Recent data suggest that the gut m(crob(ota plays a key role (n development and progress(on of 
T2DM. Th(s complex ecosystem of at least 1014 d(fferent bacter(a affects energy homeostas(s 
through the m(croorgan(sm-gut-bra(n ax(s [5, 6]. Gut m(crob(ota changes can alter 
enteroendocr(ne s(gnals and gut hormones that regulate β-cell funct(on, (nsul(n secret(on, and 
energy homeostas(s [7]. The gut m(crob(ota can affect the host's (nflammat(on response and 
energy metabol(sm, (n other words, alterat(on of the gut m(crob(ota can affect glucose and l(p(d 
metabol(sm and (nsul(n act(on. One of the most effect(ve methods of ma(nta(n(ng the balance 
of the gut m(crob(ota (s the use of prob(ot(cs, def(ned as l(ve m(croorgan(sms that, when g(ven 
(n adequate amounts, show host-spec(f(c benef(t [8]. Recently, a grow(ng number of stud(es 
have found that prob(ot(cs can alter gut flora, (mprove total cholesterol (Total-C) and Low-
Dens(ty L(poprote(n Cholesterol (LDL-C) levels [9–11], and reduce blood glucose levels and 
(nsul(n res(stance [12, 13]. Nowadays, ox(dat(ve stress (s also suggested to be a mechan(sm 
underly(ng d(abetes and (ts compl(cat(ons. Ant(ox(dant mechan(sms of prob(ot(cs (nclude 
scaveng(ng of react(ve oxygen spec(es, metal (on chelat(on, enzyme (nh(b(t(on, reduct(on 
act(v(ty, and (nh(b(t(on of ascorbate autox(dat(on [14].  
Mod(f(cat(on of gut m(croflora w(th prob(ot(cs may be seen as a novel tool for the regulat(on of 
glucose metabol(sm and (mprovement of ox(dat(ve stress (n T2DM. Increased sever(ty of beta 
cell dysfunct(on was pos(t(vely correlated w(th (ncreased h(gh-sens(t(v(ty C-React(ve Prote(n 
(hs-CRP) concentrat(ons [15]. 
Although T2DM (s recogn(zed as a chron(c (nflammatory d(sease, the role of prob(ot(cs and 
preb(ot(cs (n (ts management rema(ns (nsuff(c(ently explored, and ex(st(ng ev(dence regard(ng 
the(r overall eff(cacy (n health and d(sease (s (ncons(stent [16].  
Regulat(on of gut m(crob(ota through d(etary (ntervent(ons (e.g., prob(ot(c (ntake) may be 
benef(c(al (n reduc(ng (nflammat(on [hs-CRP and Ceruloplasm(n (Cp)] and ox(dat(ve stress 
[Glutath(one (GSH) and Malond(aldehyde (MDA)], as well as regulat(ng glucose and l(p(d 
metabol(sm (n T2DM. The a(m of th(s study was to (nvest(gate the effects of prob(ot(cs on 
glucose and l(p(d metabol(sm, (nflammat(on and ant(ox(dant system (n pat(ents w(th T2DM. 
Methods 

Tr1al des1gn  
Th(s was a s(ngle center, prospect(ve controlled study conducted at the Internal Med(c(ne 
Outpat(ent Cl(n(c of Ege Un(vers(ty Faculty of Med(c(ne between July 2020 and June 2023. 
The study (ncluded a screen(ng v(s(t, a basel(ne v(s(t (D0) and one follow-up v(s(t dur(ng a 60-
days supplementat(on per(od. The pr(mary outcomes of the study were def(ned as the effects of 
the (ntervent(on on (nflammatory markers, spec(f(cally focus(ng on a s(gn(f(cant decrease (n 
hsCRP and Cp levels, and ox(dat(ve stress markers, (nclud(ng a s(gn(f(cant reduct(on (n MDA 
levels and a s(gn(f(cant (ncrease (n reduced GSH levels. Secondary outcomes (ncluded assess(ng 
the metabol(c and l(p(d prof(le changes, wh(ch spec(f(cally targeted a s(gn(f(cant decrease (n 
Fast(ng Blood Glucose (FBG), Postprand(al Blood Glucose (PPBG), and Glycated Hemoglob(n 
(HbA1c), alongs(de a reduct(on (n LDL-C and Tr(glycer(de (TG) levels, and an (ncrease (n 
H(gh-Dens(ty L(poprote(n Cholesterol (HDL-C) levels. 
Tr1al part1c1pants 
Power analys(s was performed and the sample s(ze was calculated as 79 w(th a stat(st(cal power 
of 80% and an effect s(ze of 0.35. S(nce two groups were to be stud(ed, 80 pat(ents were planned 
to part(c(pate (n the study to equal(ze the number of part(c(pants. 
Th(s study (ncluded adults aged 35-65 years who were d(agnosed w(th T2DM accord(ng to 
Amer(can D(abetes Assoc(at(on gu(del(nes, who volunteered to part(c(pate (n th(s study. 
Exclus(on cr(ter(a were as follows: Use of any system(c ant(b(ot(cs, mult(v(tam(ns, m(nerals, 
herbal med(c(nes, preb(ot(c, prob(ot(c and postb(ot(c supplements (n the last 3-6 months, any 
(nflammatory 
bowel 
d(sease, 
severe 
renal 
dysfunct(on 
or 
hepat(c 
dysfunct(on, 
(mmunodef(c(ency d(seases, acute (nfect(on, rheumato(d arthr(t(s, cancer h(story, h(story of 
alcohol abuse or drug dependence, pregnant or lactat(ng women. D(etary hab(ts, phys(cal 
act(v(ty, and glucose-lower(ng therapy of the part(c(pants were not (ntervened dur(ng the study.  
Part(c(pants were allocated sequent(ally accord(ng to order of presentat(on (alternate 
allocat(on). Accord(ng to the order of presentat(on to the outpat(ent cl(n(c, the f(rst pat(ent who 
met the (nclus(on cr(ter(a was (ncluded (n the prob(ot(c group and the second pat(ent who 
presented to the outpat(ent cl(n(c was (ncluded (n the control group. Pat(ents (n the (ntervent(on 
group were g(ven a prob(ot(c supplement conta(n(ng Lactobac'llus ac'doph'lus, Lactobac'llus 
rhamnosus, B'f'dobacter'um lact's and Lactobac'llus paracase', each conta(n(ng 1.25 b(ll(on 
l(ve m(croorgan(sms, w(thout v(tam(ns and m(nerals, tw(ce a day (n add(t(on to the(r current 
treatment for 12 weeks (Prob(ot(c group). Pat(ents (n the control group were not g(ven any 

add(t(onal treatment and were allowed to cont(nue the(r ant(d(abet(c treatment for 12 weeks 
(Control group). Part(c(pants tak(ng prob(ot(cs were (nstructed to d(scont(nue prob(ot(c (ntake 
(f they exper(enced any ser(ous adverse events requ(r(ng (ntervent(on. The prob(ot(c product 
was suppl(ed w(thout placebo equ(valent, and due to COVID-19-related supply l(m(tat(ons no 
(dent(cal-appear(ng placebo powder could be obta(ned. Soc(odemograph(c data and med(cal 
data were recorded for all part(c(pants. 
Anthropometr1c Measurements  
Body we(ght, he(ght, wa(st (WC) and h(p c(rcumferences (HC) were collected follow(ng the 
standard(zed procedures recommended (n the World Health Organ(zat(on Expert Comm(ttee 
Gu(del(nes for phys(cal exam(nat(on at basel(ne and after 12 weeks [17]. Body mass (ndex 
(BMI) (kg/m2) and wa(st-to-h(p rat(o (WHR) (WC/HC) were calculated. 
A Food Frequency Quest1onna1re (FFQ)  
A Food Frequency Quest(onna(re was employed to determ(ne how often spec(f(c food groups 
or (nd(v(dual food (tems were consumed. The frequency of consumpt(on was recorded on a 
da(ly, weekly, b(weekly, or monthly bas(s, prov(d(ng (ns(ght (nto part(c(pants’ overall d(etary 
patterns. The FFQ (s a commonly used tool to assess the relat(onsh(p between d(etary hab(ts 
and health outcomes and can be adapted (n var(ous ways depend(ng on the researcher’s 
object(ves. Food (tems can be l(sted (nd(v(dually or grouped accord(ng to categor(es such as 
full-fat, low-fat, or fat-free opt(ons [18]. The FFQ cons(sted of 68 (nd(v(dual food (tems 
clustered (nto f(ve ma(n food groups. In th(s study, the FFQ (ncluded f(ve ma(n food groups: 
da(ry products, meat-egg-legumes, fru(ts and vegetables, cereals, fats and sweets, and 
beverages. Part(c(pants were asked to (nd(cate how often they consumed each (tem (e.g., 1-2 
t(mes da(ly, 1-3 t(mes weekly, 4-6 t(mes weekly, once every two weeks, once monthly, or never), 
wh(ch enabled the calculat(on of the(r food consumpt(on frequency over the past month. 
The Internat1onal Phys1cal Act1v1ty Quest1onna1re (IPAQ)  
When apply(ng the IPAQ, part(c(pants were asked about the(r v(gorous phys(cal act(v(ty, 
moderate phys(cal act(v(ty, and walk(ng durat(on (n the last seven days. Add(t(onally, the 
number of days per week they performed these act(v(t(es and t(me spent s(tt(ng were determ(ned 
[19]. The Turk(sh val(d(ty and rel(ab(l(ty study of the quest(onna(re was conducted by Öztürk 
[20]. Scor(ng was calculated as Metabol(c Equ(valent of Task (MET) scores from the sum of 
durat(on and frequency of moderate act(v(ty, v(gorous act(v(ty, and walk(ng [21]. Categor(es: 
• Category I: Inact(ve (nd(v(duals (<600 MET-m(n/week) 

• Category II: M(n(mum act(ve (nd(v(duals (600-3000 MET-m(n/week) 
• Category III: Very act(ve (nd(v(duals (>3000 MET-m(n/week) 
Ferrans and Powers Qual1ty of L1fe Index© D1abetes Vers1on- III 
The scale was developed by Ferrans and Powers to measure qual(ty of l(fe (n d(abet(c pat(ents, 
cons(st(ng of 34 sat(sfact(on and 34 (mportance (tems. It (ncludes 5 subscales: Total qual(ty of 
l(fe, Health and Funct(on(ng, Soc(al and Econom(c, Psycholog(cal/Sp(r(tual, and Fam(ly scores 
[22]. H(gher scores (nd(cate better qual(ty of l(fe. The Turk(sh val(d(ty study was conducted by 
Ozer [23].  
Eat1ng Att1tudes Test (EAT-26)  
The EAT-26 was developed for Garner et al.[24].  The test cons(sts of 26 (tems, and the total 
score (s between 0-53. A score of 20 and above (s def(ned as “abnormal eat(ng behav(or” and a 
score less than 20 (s def(ned as “normal eat(ng behav(or”. The Turk(sh val(d(ty and rel(ab(l(ty 
study was conducted by Erguney-Okumus and Sertel-Berk [25]. 
Biochemical measurements 
Fasting blood samples were collected from the participants at baseline and week 12, following 
overnight fasting and after 2 hours of satiety. FBG, PPBG, HbA1c, Total-C, LDL-C, HDL-C, 
TG and hs-CRP were measured using an autoanalyzer. Serum MDA levels were determined by 
the thiobarbituric acid method [26], GSH levels by enzymatic recycling method [27] and Cp 
levels by colorimetric method [27] using a microplate reader. 
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