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Biomedical subjects

Rakesh K Sharma

Publications and source records attributed to Rakesh K Sharma.

At least 19 recordsLinked to original sources
PubMed2006

Automation is the key to standardized semen analysis using the automated SQA-V sperm quality analyzer.

OBJECTIVE: To evaluate the performance of the automated semen quality analyzer system for assessing sperm quality. DESIGN: Double-blind prospective study. SETTING: Tertiary care hospital. PATIENT(S): Fifty healthy men donated semen samples. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Precision, accuracy and agreement between automated and manual semen analysis methods was assessed for sperm concentration, motility, morphology, and known concentrations of latex bead quality control media. RESULT(S): A good agreement was seen between the results of sperm concentration reported by the SQA-V automated analyzer (Spermalite/SQA-V; Medical Electronic Systems Ltd, Caesarea Industrial Park, Israel) and those obtained manually. A similar linearity was seen when the SQA-V results were compared with the manual data and also when the manual results of individual operators were compared with each other. The automated assessment of morphology showed high sensitivity (89.9%) for identifying percent normal morphology, and the precision of the SQA-V was considerably higher when compared with the manual method. The interoperator variability for manual assessment was significant. The automated analysis was quick compared with the manual method. CONCLUSION(S): The SQA-V can be used interchangeably with manual semen analysis methods for examining sperm concentration and motility. The automated SQA-V analyzer is more precise and shows the ability to accurately classify normal versus abnormal sperm morphology.

Diagnosis, Computer-Assisted↗
PubMed2006

Biological evaluation, chelation, and molecular modeling studies of novel metal-chelating inhibitors of NF-kappaB-DNA binding: structure activity relationships.

Previously, we have reported that aurintricarboxylic acid (ATA) is one of the most potent inhibitors of the DNA binding of transcription factor NF-kappaB. We now report the NF-kappaB-DNA binding inhibitory activity of ATA analogues. An electrophoretic mobility shift assay has shown that bromopyrogallol red (BPR) is the most effective inhibitor of NF-kappaB-DNA binding among the studied analogues. The molecular modeling studies showed that BPR makes a strong network of hydrogen bonds with the DNA-binding region of the p50 subunit of NF-kappaB and has electronegative potential on its peripheral surface. Because zinc has been reported to influence the DNA binding of NF-kappaB, the interaction of these analogues with zinc was studied. Chemical speciation and formation-constant studies showed that BPR forms the most stable 1:1 complex with zinc. BPR has also been found to be the most potent antioxidant among the studied analogues.

Antiviral Agents↗
PubMed2006

Role of total antioxidant capacity in the differential growth of human embryos in vitro.

OBJECTIVE: The objective of this study was to examine the relationship of early human embryonic development parameters with day 1 culture media total antioxidant levels (day 1 TAC). DESIGN: Prospective study. SETTING: Patients undergoing assisted reproduction (ART) in a teaching hospital. PATIENT(S): Patients undergoing conventional IVF (n = 153; 167 cycles) and intracytoplasmic sperm injection (ICSI; n = 105; 116 cycles) were included. Both fertilization and early cultures were performed in human tubal fluid (HTF) with 5% serum substitute supplement. INTERVENTION(S): Levels of total antioxidants in the central well (sample) and the outer well (control) of each embryo culture dish were measured. MAIN OUTCOME MEASURE(S): Fertilization rate and embryo quality at days 3 and 5 were recorded for each cycle. Age, parity, and demographic features were also compared. RESULT(S): After controlling for all demographic and clinical variables, day 1 TAC was related to fertilization rates in both groups of patients. Day 1 TAC was significantly related to high (>7) day 3 cell number, low (<10%) fragmentation rate, and blastocyst development rate in both conventional IVF and ICSI cycles. Day 1 TAC was related to pregnancy in ICSI but not conventional IVF cycles. CONCLUSION(S): Day1 TAC appears to be an important biochemical marker for early embryonic growth. Decreased embryonic fragmentation, enhanced cleavage rate, and increased blastocyst development rate may be partially related to day 1 TAC in the culture media. Whether this relationship is a cause or effect needs further assessment.

Adult↗
PubMed2006

Significance of sperm characteristics in the evaluation of male infertility.

OBJECTIVE: To compare sperm characteristics among: patients undergoing infertility evaluation, patients with male factor infertility (MFI), healthy sperm donors, and men with proven fertility; to examine the overlap of sperm characteristics in all four of these groups; and to identify good discriminators of fertility versus infertility among sperm characteristics. DESIGN: Retrospective study. SETTING: Male infertility clinic at a tertiary care hospital. PATIENT(S): Proven fertile men (n = 56), normal donors (n = 91), men presenting for infertility evaluation (n = 406), and MFI patients (n = 166). INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Routine semen analysis. RESULT(S): Using current World Health Organization (WHO) reference values, a large group of MFI patients presented with higher sperm concentration (27.5 x 10(6) to 99.2 x 10(6)), resulting in broader overlap with fertile men and poor sensitivity (0.48). Similarly, percentage normal morphology (%) using Tygerberg's strict criteria was low in most of the MFI patients (sensitivity 0.83), almost half of the fertile men also presented with abnormal morphology (specificity 0.51). Of all the variables examined, sperm motility (%) was superior, having minimum overlap range (lower and upper cut-off values 46% and 75%) and high sensitivity (0.74) and specificity (0.90). Areas under curve were higher for motility (0.90) and concentration (0.84) compared with morphology (WHO 0.65 and Tygerberg's strict criteria 0.74). CONCLUSION(S): Sperm motility and concentration provide more accurate information than morphology (WHO and Tygerberg's criteria) during infertility evaluation. Redefining the reference values for concentration and morphology may significantly increase the importance of routine semen analysis.

Case-Control Studies↗
PubMed2006

Reactive oxygen species as an independent marker of male factor infertility.

OBJECTIVE: To determine the abnormal patterns of reactive oxygen species (ROS) production in male factor infertility (MFI) patients and to define the ROS reference values in such patients. DESIGN: A retrospective study. SETTING: Male infertility clinic at a tertiary healthcare center. PATIENT(S): We examined 132 MFI patients (all normal sperm parameters, n = 24, and all abnormal sperm parameters, n = 38) and 34 healthy donors. INTERVENTION(S): Routine semen analysis, measurement of ROS. MAIN OUTCOME MEASURE(S): Sperm parameters, ROS levels (10(4) cpm/20 x 10(6) sperm). RESULT(S): Normal, healthy donors had significantly higher (P<.0001) sperm concentration, motility, and morphology compared with all MFI patients. Univariate analysis indicated a significant association between MFI and log (ROS + 1) (odds ratio [OR] = 3.84), besides sperm parameters and age. A multivariate model using logistic regression analysis also indicated an independent association of log ROS with MFI (OR = 4.25). The ROS cutoff values of 1.2-1.4 had a sensitivity of 0.70-0.78 with a corresponding specificity of 0.82. However, at a cutoff point of 1.2, the OR was 68.6, which increased with an increase in the cutoff. CONCLUSION(S): High ROS is an independent marker of MFI, irrespective of whether these patients have normal or abnormal semen parameters. We suggest the inclusion of ROS measurement as part of idiopathic infertility evaluation. Treatment with antioxidants may be beneficial in such patients.

Adult↗
PubMed2006

Cobalt(II) complexes of biologically active glutathione: spectroscopic and molecular modeling studies.

Cobalt(II) complexes of reduced glutathione (GSH) of general composition Na[Co(L)(X)].nH2O (where H2L = GSH; X = Cl-, NO3-, NCS-, CH3CO2-, HCO2-, ClO4- and n = 0-4) have been synthesized and characterised by elemental analyses, vibrational spectra, electronic spectra, magnetic susceptibility measurements, thermal studies and molecular modeling studies. Electronic spectra indicate planar geometry for all the complexes. Infrared spectra indicate the presence of H2O molecules (except perchlorate complex) in the complexes that has been supported by TG/DTA. The room temperature magnetic moment values for all complexes lie in the range of 2.60-2.80 BM range indicating departure from spin only values due to second order Zeeman effect. Thermal decomposition of all the complexes proceeds via first order kinetics. The Na[Co(L)(Cl)].2H2O complex has the minimum activation energy and Na[Co(L)(CH3CO2)].3H2O has the maximum activation energy. The molecular modeling calculation for energy minimization optimizes geometry of the metal complexes.

Cobalt↗
PubMed2006

Delivery of hydrophobised 5-fluorouracil derivative to brain tissue through intravenous route using surface modified nanogels.

Random copolymeric micelles composed of N-isopropylacrylamide (NIPAAM) and N-vinylpyrrolidone (VP) cross-linked with N,N'-methylenebisacrylamide (MBA) have been used as nanogel carriers to encapsulate N-hexylcarbamoyl-5-fluorouracil (HCFU), a prodrug of 5-FU, and have been targeted to brain tissue across blood-brain barrier (BBB) after coating with polysorbate 80. Accumulation of nanogel particles in the brain and other tissues of "strain A" mice had been monitored by radiolabeling of nanogels with (99m)Tc. Gamma Scintigraphic technique was also performed to see the distribution of (99m)Tc labeled nanogels in the brain. The retention time in blood appeared to be slightly longer for coated nanogels than that of uncoated nanogels though the accumulation of coated nanogels in the RES was more or less same as that of uncoated nanogels. The blood however had almost double accumulation of polysorbate 80 coated nanogels in the initial 5 min compared to that shown by uncoated nanogels. We speculate that coating of nanogels with polysorbate 80 alters the surface properties of nanogels, which results in relatively higher uptake in the brain tissue. The studies revealed that a large portion of (99m)Tc labeled HCFU loaded nanogels are accumulated in the RES (lung, liver and spleen). The accumulation of the labeled nanogels in the brain, however, is much less compared to RES and it has been found that while an amount of uncoated labeled nanogels was found to be 0.18% of the injected dose, it increased to 0.52% on coating with polysorbate 80. The optimal amount of polysorbate 80 added to nanogels for the maximum delivery of particles to brain was found to be 1% w/w. These results were further supported by the gamma scintigrams of New Zealand rabbits. Thus, the present nanogel system has opened a new avenue for poorly soluble drugs to be targeted to brain by coating the particles with polysorbate 80.

Animals↗
PubMed2006

Hematopoietic stem cell antigen CD34: role in adhesion or homing.

CD34 is highly glycosylated surface antigen of enormous clinical utility in the identification, enumeration, and purification of engraftable lymphohematopoietic progenitors for transplantation. However, recently its importance in the specific marking of most immature hematopoietic stem/progenitor cells have been questioned by addressing long-term reconstitution capability of CD34(-) hematopoietic cellular fractions. These controversies have stimulated a demand for elucidation of the structure, function, and molecular interactions of CD34 to define exactly its biological significance in clinical regimens. There is accumulating data showing the participation of CD34 in adhesion or perhaps homing of lymphohematopoietic progenitors. On the other hand, CD34 has been demonstrated to down-regulate cytokine-induced differentiation and proliferation of CD34(+) cells. Studies in CD34 knockout mice revealed normal hematopoiesis but a profound delay in hematopoietic reconstitution after sublethal irradiation of the mice. In short, CD34 expression is likely to represent a specific state of hematopoietic development that may have altered adhering properties with expanding and differentiating capabilities in both in vitro and in vivo conditions. This article focuses on the adhesive properties of CD34 and its potential role in homing, which are likely to mimic lymphocyte homing to the inflammatory sites.

Animals↗
PubMed2005

Peritoneal fluid leptin is associated with chronic pelvic pain but not infertility in endometriosis patients.

BACKGROUND: Leptin influences the proinflammatory immune responses and has angiogenic activity in vitro and in vivo. The objective of this study was to evaluate the peritoneal fluid levels of leptin in patients with endometriosis and idiopathic infertility and compare them with a control group of tubal ligation/reanastomosis patients. METHODS: In this observational, prospective controlled study, peritoneal fluid from 108 women was obtained while they underwent laparoscopy for pelvic pain, infertility, tubal ligation or sterilization reversal. We measured the concentration of leptin in the peritoneal fluid and compared the levels among women who were divided into groups according to their post-surgical diagnosis. Sixty patients were diagnosed with endometriosis, 10 with idiopathic infertility and 38 had undergone tubal ligation or reanastomosis (control group). RESULTS: Peritoneal fluid leptin was significantly higher in endometriosis 14.62+/-9.79 (mean+/-SD) ng/ml compared to idiopathic infertility [0.92+/-1.57 ng/ml (P=0.0007)] and to controls [0.78+/-1.94 ng/ml (P<0.0001)]. Leptin levels were positively correlated with the stage of endometriosis (r=0.45; P=0.03), and with pelvic pain in endometriosis patients (r=0.49; P=0.001). Peritoneal fluid leptin levels in patients with idiopathic infertility were comparable to controls. CONCLUSIONS: Higher levels of leptin were observed in peritoneal fluid of patients with endometriosis compared to those without the disease. These data suggest that the proinflammatory and neoangiogenic action of leptin may contribute to the pathogenesis of endometriosis. Moreover, leptin may play a role in endometriosis-associated pain.

Ascitic Fluid↗
PubMed2005

Identification of male factor infertility using a novel semen quality score and reactive oxygen species levels.

PURPOSE: To determine whether patients with male factor infertility can be accurately identified by calculating a novel semen quality score and measuring levels of reactive oxygen species during routine infertility screening. METHODS: Semen samples from 133 patients and 91 healthy donors were evaluated with manual and computer-assisted semen analysis. A principal component analysis model was employed to calculate a semen quality score. In brief, this score was calculated by base 10 logarithms multiplied by varying weights given to 9 sperm parameters. Reactive oxygen species levels were measured using chemiluminescence assay. RESULTS: The semen quality score had a sensitivity of 80.45% and accuracy of 77% at a cutoff of 93.1 in identifying patients with male factor infertility. The area under the receiver operating characteristic curves for the semen quality score was 84.28% (95% CI: 65.22%-100%). Reactive oxygen species levels [log10 (reactive oxygen species +1)] were significantly higher in male factor infertility patients. Reactive oxygen species had a sensitivity of 83.47% and specificity of 60.52% with an accuracy of 75% at a cutoff of 1.25 in identifying male factor infertility patients. The area under the receiver operating characteristic curve for reactive oxygen species levels was 78.92% (95% CI: 72.60%-85.23%). Semen quality scores were significantly and negatively correlated with reactive oxygen species levels in the donors and the male factor infertility patients. CONCLUSIONS: The semen quality score and reactive oxygen species levels in semen samples appear to be strongly associated with male factor infertility. Because both of these parameters are more sensitive than individual sperm parameters in identifying male factor infertility, they should be included in routine infertility screening.

Case-Control Studies↗
PubMed2005

Role of oxidative stress in female reproduction.

In a healthy body, ROS (reactive oxygen species) and antioxidants remain in balance. When the balance is disrupted towards an overabundance of ROS, oxidative stress (OS) occurs. OS influences the entire reproductive lifespan of a woman and even thereafter (i.e. menopause). OS results from an imbalance between prooxidants (free radical species) and the body's scavenging ability (antioxidants). ROS are a double-edged sword - they serve as key signal molecules in physiological processes but also have a role in pathological processes involving the female reproductive tract. ROS affect multiple physiological processes from oocyte maturation to fertilization, embryo development and pregnancy. It has been suggested that OS modulates the age-related decline in fertility. It plays a role during pregnancy and normal parturition and in initiation of preterm labor. Most ovarian cancers appear in the surface epithelium, and repetitive ovulation has been thought to be a causative factor. Ovulation-induced oxidative base damage and damage to DNA of the ovarian epithelium can be prevented by antioxidants. There is growing literature on the effects of OS in female reproduction with involvement in the pathophysiology of preeclampsia, hydatidiform mole, free radical-induced birth defects and other situations such as abortions. Numerous studies have shown that OS plays a role in the pathophysiology of infertility and assisted fertility. There is some evidence of its role in endometriosis, tubal and peritoneal factor infertility and unexplained infertility. This article reviews the role OS plays in normal cycling ovaries, follicular development and cyclical endometrial changes. It also discusses OS-related female infertility and how it influences the outcomes of assisted reproductive techniques. The review comprehensively explores the literature for evidence of the role of oxidative stress in conditions such as abortions, preeclampsia, hydatidiform mole, fetal embryopathies, preterm labour and preeclampsia and gestational diabetes. The review also addresses the growing literature on the role of nitric oxide species in female reproduction. The involvement of nitric oxide species in regulation of endometrial and ovarian function, etiopathogenesis of endometriosis, and maintenance of uterine quiescence, initiation of labour and ripening of cervix at parturition is discussed. Complex interplay between cytokines and oxidative stress in the etiology of female reproductive disorders is discussed. Oxidant status of the cell modulates angiogenesis, which is critical for follicular growth, corpus luteum formation endometrial differentiation and embryonic growth is also highlighted in the review. Strategies to overcome oxidative stress and enhance fertility, both natural and assisted are delineated. Early interventions being investigated for prevention of preeclampsia are enumerated. Trials investigating combination intervention strategy of vitamin E and vitamin C supplementation in preventing preeclampsia are highlighted. Antioxidants are powerful and there are few trials investigating antioxidant supplementation in female reproduction. However, before clinicians recommend antioxidants, randomized controlled trials with sufficient power are necessary to prove the efficacy of antioxidant supplementation in disorders of female reproduction. Serial measurement of oxidative stress biomarkers in longitudinal studies may help delineate the etiology of some of the diosorders in female reproduction such as preeclampsia.

Aging↗
PubMed2005

Studies on structure activity relationship of some dihydroxy-4-methylcoumarin antioxidants based on their interaction with Fe(III) and ADP.

Three dihydroxy-4-methylcoumarin (DHMC) derivatives, namely 7,8-DHMC, 6,7-DHMC and 5,7-DHMC alone and complexed with Fe(III) and ADP have been tested for their antioxidative potential. Chemical speciation studies and formation constants reveal the formation of strong DHMC-Fe-ADP (1:1:1) ternary complex. In vitro studies were done for their antioxidative property by scavenging the superoxide radicals (O2*-) generated by xanthine + xanthine oxidase (XO) reaction. The IC50 values for 7,8-DHMC, 6,7-DHMC and 5,7-DHMC and their ternary complexes with Fe(III)-ADP worked out to be 34.0 microM, 62.0 microM, 8.80 mM and 10.5, 11.5 and 148.5 microM, respectively. The results indicate that O2*- scavenging potential of all the three DHMCs increased significantly after forming the ternary complex with Fe(III) and ADP. The structure activity relationship studies suggest that the introduction of hydroxyl group at 7th and 8th positions in the coumarins, irrespective of Fe(III)-ADP complexation, increases the antioxidative efficacy. No change in uric acid production in the reactions done for all studies further reveals that the coumarin derivatives and their complexes were the only causative factors for O2*- scavenging and not the suppression of the enzyme, xanthine oxidase.

Adenosine Diphosphate↗
PubMed2005

Effect of pentoxifylline in reducing oxidative stress-induced embryotoxicity.

PURPOSE: To 1) evaluate the embryotoxic effects of hydrogen peroxide on mouse embryo development and 2) examine if pentoxifylline can reverse hydrogen peroxide induced embryotoxicity. METHODS: Prospective in vitro study examining the effects of varying concentrations of hydrogen peroxide and pentoxifylline on the blastocyst development rate alone as well as in combination. RESULTS: A dose-dependent decrease in % BDR was seen with increasing concentrations of H2O2. High concentrations of hydrogen peroxide (> 60 microM) were embryotoxic. Pentoxifylline (500 microM) was able to reduce the embryotoxic effect of hydrogen peroxide. Percent blastocyst development rate increased from 44% in hydrogen peroxide alone to 85% in hydrogen peroxide and pentoxifylline coincubation. CONCLUSIONS: Pentoxifylline may be beneficial in reducing hydrogen peroxide induced embryo damage and improve IVF outcome. Patients with endometriosis-associated infertility may benefit from the use of pentoxifylline without significantly affecting embryo development.

Animals↗
PubMed2005

Inhibitory activity of polyhydroxycarboxylate chelators against recombinant NF-kappaB p50 protein-DNA binding.

The inhibitory effect of 7,8-dihydroxy-4-methylcoumarin (7,8-DHMC), 5,7-dihydroxy-4-methylcoumarin (5,7-DHMC), and gallic acid on the DNA binding of recombinant p50 protein and their interaction with zinc ion were studied. Electrophoretic mobility shift assay (EMSA) using p50 and biotin labeled DNA has shown that gallic acid is more effective than the dihydroxycoumarins in inhibiting the p50-DNA binding. Molecular modeling studies suggest an explanation for these observations. Effect of the addition of zinc after p50-DNA-binding inhibition by gallic acid was also studied. Chemical speciation and formation constant studies show that gallic acid forms a more stable 1:1 complex with zinc ion in comparison to the dihydroxycoumarins.

Cations, Divalent↗
PubMed2005

Characterization of oxidative stress status by evaluation of reactive oxygen species levels in whole semen and isolated spermatozoa.

We defined the basal levels of reactive oxygen species (ROS) in normal donors in neat (whole unprocessed) semen specimens, and in mature and immature spermatozoa isolated by a double-density gradient technique. In addition, we demonstrated that the ROS levels were significantly lower in neat semen compared with washed spermatozoa. The reference values of ROS in neat semen and mature spermatozoa can be used to define the pathologic levels of ROS in infertile men and may guide in therapeutic interventions.

Adult↗
PubMed2005

Impact of sperm morphology on DNA damage caused by oxidative stress induced by beta-nicotinamide adenine dinucleotide phosphate.

OBJECTIVE: To investigate the role of DNA damage induced by beta-nicotinamide adenine dinucleotide phosphate (NADPH) in human spermatozoa. DESIGN: Prospective controlled study. SETTING: Male infertility clinic at the Glickman Urological Institute, Cleveland Clinic Foundation, Cleveland, Ohio. PATIENT(S): Twenty-eight men undergoing infertility screening. INTERVENTION(S): Chemiluminescence assay and terminal deoxynucleotidyl transferase-mediated digoxigenin-dUTP nick-end labeling (TUNEL) assay coupled flow cytometry after incubating mature and immature sperm separated by density gradient with 5 mM NADPH for 0, 3, and 24 hours. MAIN OUTCOME MEASURE(S): Reactive oxygen species (ROS) generation (10(6) counted photons per minute/10(6) sperm) and percentage of spermatozoa with fragmented DNA. RESULT(S): Immature sperm from teratozoospermic semen samples were characterized by a statistically significant presence of cytoplasmic residues in the mid-piece when compared with mature normozoospermic samples. Increased ROS production was observed in spermatozoa rich in cytoplasmic residues that showed a statistically significant positive correlation with sperm DNA damage in a time-dependent manner. CONCLUSION(S): Immature sperm contain high nicotinamide adenine dinucleotide phosphate (NADPH) in cytoplasmic droplets, but it has not yet been clear whether abnormal sperm morphology plays any role in sperm DNA damage induced by oxidative stress. Our data support the role of NAPDH in ROS-mediated sperm DNA damage and suggest that abnormal sperm morphology combined with elevated ROS production may serve as a useful indicator of potential damage to sperm DNA.

DNA Damage↗
PubMed2005

Infliximab may reverse the toxic effects induced by tumor necrosis factor alpha in human spermatozoa: an in vitro model.

OBJECTIVE: To examine the toxic effects of tumor necrosis factor alpha (TNF-alpha) on ejaculated spermatozoa and evaluate the ability of infliximab to reverse these effects. DESIGN: Prospective controlled study. SETTING: Cleveland Clinic Foundation, Cleveland, Ohio. PATIENT(S): Thirty-one healthy sperm donors. INTERVENTION(S): Exposure of human spermatozoa to varying concentrations of TNF-alpha (100, 300, 400, 500 pg/mL, and 2.5 microg/mL) and infliximab (400 microg/mL). MAIN OUTCOME MEASURE(S): Sperm motility, functional integrity of plasma membrane, and DNA fragmentation. RESULT(S): Spermatozoa quality declined following incubation with TNF-alpha in a dose-dependent and time-dependent manner. Sperm motility and membrane integrity were higher in the samples incubated with TNF-alpha plus infliximab than in the samples treated with TNF-alpha only. These parameters improved significantly and were comparable with both controls and sperm incubated with infliximab alone. Similarly, the percentage of spermatozoa with DNA fragmentation improved significantly following incubation with TNF-alpha plus infliximab and again was comparable with both controls and sperm incubated with infliximab alone. CONCLUSION(S): Spermatozoa may be exposed to abnormal levels of TNF-alpha in the male reproductive tract or during their passage into the female reproductive tract (in cases of endometriosis). Exposing spermatozoa to pathological concentrations of TNF-alpha can result in significant loss of their functional and genomic integrity. Infliximab could potentially be used to help treat female infertility caused by endometriosis in those with elevated levels of TNF-alpha in their peritoneal fluid.

Analysis of Variance↗