Evaluation of Circulating Leptin and Its Receptor (Ob-R) Tissue Expression in Colorectal Cancer, a Report From North of Iran.

Background & Objective: Leptin is an adipocyte-derived hormone with a critical role in energy balance. As demonstrated by previous investigations, leptin acts as a proliferative and angiogenic factor in cancer cells. However, results regarding its role in colorectal cancer are still inconclusive. We aimed to evaluate serum leptin and tissue expression of leptin receptor (Ob-R) in normal and malignant samples of colorectal. Methods: Serum and tissue samples from pathology-confirmed colorectal cancer patients and normal controls referring to a university hospital of Mazandaran were obtained during 2019-21. ELISA and immunohistochemistry were applied to determine leptin and Ob-R expression respectively. Results: A total of 90 samples belonging to 46 normal and 44 CRC patients were enrolled. Normal and CRC groups included 32 (69.56%) and 21 (47.72%) female subjects respectively. The average leptin concentration in the normal group was 115.80 and, in the patient, group was 124.47 ng/mL (P=0.897). CRC cases showed an insignificantly higher Ob-R detection rate (P=0.086). Conclusion: There was no significant difference in leptin and Ob-R expression between CRC patients and normal subjects. Thus, leptin and its receptor may not be useful as a biomarker of CRC.

Dopamine receptors gene overexpression in the microenvironment of invasive gastric cancer and its potential implications.

BACKGROUND: Gastric cancer (GC) is the fifth most common cancer worldwide and the most commonly diagnosed cancer in Iran. The nervous system provides proximity to tumor cells by releasing neurotransmitters such as dopamine and presenting them to the corresponding receptor-bearing tumors. While nerve fibers infiltrate the tumor microenvironment, little is known about the expression levels of dopamine (DA), dopamine receptors (DRs), and catechol-O-methyltransferase (COMT) in GC patients. METHODS: DRs and COMT expression were analyzed in 45 peripheral blood mononuclear cells (PBMCs) and 20 paired tumor and adjacent tissue of GC patients by quantitative polymerase chain reaction. DA was measured in plasma specimens using enzyme-linked immunosorbent assay. Protein-protein interaction analysis was carried out to identify GC-related hub genes. RESULTS: Increased expression of DRD1-DRD3 was found in tumor specimens compared with adjacent non-cancerous specimens (P < 0.05). A positive correlation was found between DRD1 and DRD3 expression (P = 0.009); DRD2 and DRD3 expression (P = 0.04). Plasma levels of dopamine were significantly lower in patients (1298 pg/ml) than in controls (4651 pg/ml). DRD1-DRD4 and COMT were up-regulated in PBMCs of patients compared with controls (P < 0.0001). Bioinformatic analyses showed 30 hub genes associated with Protein kinase A and extracellular signal-regulated kinase signaling pathways. CONCLUSIONS: The findings indicated dysregulation of DRs and COMT mRNA expression in GC and suggest that the brain- gastrointestinal axis may mediate gastric cancer development. Network analysis revealed that combination treatments could be considered for optimizing and improving the precision treatment of GC.

Serotonin Receptors and Acetylcholinesterase Gene Expression Alternations: The Potential Value on Tumor Microenvironment of Gastric Cancer.

INTRODUCTION: Gastric cancer is one of the common causes of cancer-related death in the world. Neurotransmitters have recently been related to the proliferation of cancer cells, but the role of neurotransmitters in the progression of gastric cancer is still unexplored. The cross-talk between the nervous system and immune cells through serotonin and its receptors in the tumor microenvironment can impact tumor progress. Our purpose is to expose probable changes in serotonin receptors, acetylcholinesterase, and monoamine oxidase A gene expression in gastric cancer. METHODS: Transcript of serotonin receptors (5-HTR2A, 5-HTR2B, 5-HTR3A, 5-HTR7) and monoamine oxidase A genes in the peripheral blood mononuclear cells (40 patients and 40 control) and tissue (21 tumors and 21 normal adjacent tissues) were assessed. The gene expression was analyzed by quantitative real-time PCR using suitable primers. Statistical analysis was performed using appropriate software (REST, Prism). RESULTS: Significantly higher amounts of 5-HTR2A, 5-HTR2B, 5-HTR3A, 5-HTR7, and acetylcholinesterase gene transcripts were found in the peripheral blood of gastric cancer patients compared with healthy individuals. The expression of 5-HTR2B and 5-HTR3A genes was significantly higher (p = 0.0250, p = 0.0005, respectively) and the acetylcholinesterase gene was lower in the tissue of patients (p = 0.0119) compared with adjacent normal tissue. CONCLUSION: This study highlights the role of serotonin receptors in gastric cancer that might have suggestions for the development of novel therapeutics and defensive approaches that target factors associated with the link between the nervous system, cancer cells, and the tumor microenvironment.

Significance of Serum Apelin Levels as a Biomarker for Gastric Cancer Diagnosis.

OBJECTIVES: Gastric cancer (GC) is one of the common lethal disease and the most common cancers worldwide and in Iran. North of Iran is known as a common area of gastric cancer and a high-risk zone in Iran. Apelin is a biomolecule that plays roles in various types of cancers. This study was designed to investigate the serum apelin-12 levels in patients with gastric cancer as a predictive marker and affordable noninvasive alternative. METHODS: In this case-control study, the case group included 42 patients with gastric cancer who were diagnosed by endoscopy and pathological findings. The participants in the case group were compared with the control group including 43 healthy individuals with no history of gastric cancer in their first-degree relatives and visiting the lab for routine tests. Apelin-12 serum level was assessed using ELISA kit. Data were analyzed in SPSS V16.0 applying Fisher's exact test, Mann-Whitney U test, and t-test. RESULTS: Serum apelin-12 in patients with gastric cancer was found to be statistically lower than that in healthy individuals (p< 0.05). There were no significant differences between clinicopathological characteristics and apelin-12 expression. The median survival time in experimental and control groups was 16.0 months. CONCLUSIONS: In the current study, serum levels of apelin were significantly different between cases and controls.

Can we consider soluble herpes virus entry mediator (sHVEM) as a tumor marker?

Background: Immune checkpoint molecules have critical roles in directing immune responses into co-inhibitory and co-stimulatory signals. Herpes virus entry mediator (HVEM) is a receptor of tumor necrosis factor receptor superfamily with unique features due to its interaction with both inhibitory and stimulatory ligands. The aim of this study was to measure the serum level of the soluble form of HVEM in patients with gastric, colorectal and breast cancers and evaluating its diagnostic and prognostic value. Methods: The concentration of the soluble HVEM (sHVEM) was determined in the serum of 36 patients with breast cancer, 50 patients with colorectal cancer and 59 patients with gastric cancer using ELISA method. Moreover, 50 healthy donors (HD) as well as 31 patients with non-ulcer dyspepsia (NUD) were used as control groups. The patients' samples were obtained from the Biobank of Cancer Research Center, Mazandaran University of Medical Sciences, Sari, Iran. Results: The level of sHVEM was significantly higher in patients with gastric (P=0.001) and breast cancer (P=0.01) than in control groups (HD). The higher level of sHVEM was observed in colorectal cancer patients in comparison with HD group, although it was not significant. Moreover, the elevated level of sHVEM was shown to be higher significantly in stage III and IV compared to stage I and II in breast cancer (P=0.03). Similar finding was detected in gastric and colorectal cancers, but not to be statistically significant. Conclusion: The results of the present study suggest that the serum level of sHVEM may be considered as a promising indicator for diagnosis as well as evaluating the progression of cancers such as gastric, breast and colorectal cancers.

Association between Serum Progastrin Biomarker Level and Gastric Cancer.

BACKGROUND AND OBJECTIVE: gastric cancer is the fifth most prevalent cancer and the fourth cause of death because of cancer. In Iran, northern and northwestern regions are considered gastric cancer hot spots. Identifying serum biomarkers could be helpful in early diagnosis of patients with gastric adenocarcinoma (GAC). Increase in progastrin level has been reported in different cancers. Given the diagnostic value of this biomarker, this study aimed to determine the diagnostic role of progastrin serum biomarker in patients with gastric cancer. METHODOLOGY: In this case-control study, forty patients with gastric cancer who were diagnosed by endoscopy and pathologic findings and visited Mazandaran Comprehensive Cancer Center. The participants had received no treatment yet and entered this study. The participants in case group were compared with the control group including forty-two individuals with no history of gastrointestinal cancer in their first-degree relatives and visiting the lab for routine tests. Progastrin serum level was assessed using ELISA kit. The Kruskal-Wallis test and Mann Whitney test, both non-parametric) were used for statistical analysis and the relation between the variables was examined using Pearson's correlation coefficient at 95% confidence level in SPSS 16. FINDINGS: In this study, progastrin serum level was significantly higher in patients with gastric cancer compared with normal participants (P = 0.035). Progastrin serum level had no significant relation with tumor clinicopathologic parameters (p-value > 0.05). CONCLUSION: Increase in progastrin may be utilized as a predictive factor for gastric cancer.

Effects of Vitamin E on Doxorubicin Cytotoxicity in Human Breast Cancer Cells in Vitro.

OBJECTIVE: This study aimed to evaluate in vitro synergistic anticancer effect of doxorubicin combined with Vitamin E. METHODS: The MTT assay was utilized to assess the cytotoxicity of Vitamin E and vitamin E combined with doxorubicin and vital activities of SKBR3, MDA-MB-231, and HFF cells over a 24-hour incubation period. In addition, the antioxidant properties of these interventions and the decrease of reactive oxygen species (ROS) content caused by the treatment were evaluated. RESULTS: The antiproliferative effect of doxorubicin increased significantly in combination with vitamin E (Doxcorobicin 2µM vs. Vitamin E 120µM, P=0.000). Despite reducing cell ROS content due to vitamin E treatment, the combination of vitamin E and doxorubicin showed no significant synergistic effect (Doxcorobicin 2µM vs. Vitamin E 120µM, P=0.998). CONCLUSION: This study indicated that the doxorubicin-vitamin E treatment reduced the viability of breast cancer cells with the minimum side effects on normal cells. In addition, the high dosage of vitamin E intensified the cytotoxicity of doxorubicin.

IFI27 transcription is an early predictor for COVID-19 outcomes, a multi-cohort observational study.

Purpose: Robust biomarkers that predict disease outcomes amongst COVID-19 patients are necessary for both patient triage and resource prioritisation. Numerous candidate biomarkers have been proposed for COVID-19. However, at present, there is no consensus on the best diagnostic approach to predict outcomes in infected patients. Moreover, it is not clear whether such tools would apply to other potentially pandemic pathogens and therefore of use as stockpile for future pandemic preparedness. Methods: We conducted a multi-cohort observational study to investigate the biology and the prognostic role of interferon alpha-inducible protein 27 (IFI27) in COVID-19 patients. Results: We show that IFI27 is expressed in the respiratory tract of COVID-19 patients and elevated IFI27 expression in the lower respiratory tract is associated with the presence of a high viral load. We further demonstrate that the systemic host response, as measured by blood IFI27 expression, is associated with COVID-19 infection. For clinical outcome prediction (e.g., respiratory failure), IFI27 expression displays a high sensitivity (0.95) and specificity (0.83), outperforming other known predictors of COVID-19 outcomes. Furthermore, IFI27 is upregulated in the blood of infected patients in response to other respiratory viruses. For example, in the pandemic H1N1/09 influenza virus infection, IFI27-like genes were highly upregulated in the blood samples of severely infected patients. Conclusion: These data suggest that prognostic biomarkers targeting the family of IFI27 genes could potentially supplement conventional diagnostic tools in future virus pandemics, independent of whether such pandemics are caused by a coronavirus, an influenza virus or another as yet-to-be discovered respiratory virus.

Lymphopenia and lung complications in patients with coronavirus disease-2019 (COVID-19): A retrospective study based on clinical data.

A rapid outbreak of novel coronavirus, coronavirus disease-2019 (COVID-19), has made it a global pandemic. This study focused on the possible association between lymphopenia and computed tomography (CT) scan features and COVID-19 patient mortality. The clinical data of 596 COVID-19 patients were collected from February 2020 to September 2020. The patients' serological survey and CT scan features were retrospectively explored. The median age of the patients was 56.7 ± 16.4 years old. Lung involvement was more than 50% in 214 COVID-19 patients (35.9%). The average blood lymphocyte percentage was 20.35 ± 10.16 (normal range, 20%-50%). Although the levels of C-reactive protein (CRP) and erythrocyte sedimentation rate (ESR) were high in more than 80% of COVID-19 patients; CRP, ESR, and platelet-to-lymphocyte ratio (PLR) may not indicate the in-hospital mortality of COVID-19. Patients with severe lung involvement and lymphopenia were found to be significantly associated with increased odds of death (odds ratio, 9.24; 95% confidence interval, 4.32-19.78). These results indicated that lymphopenia < 20% along with pulmonary involvement >50% impose a multiplicative effect on the risk of mortality. The in-hospital mortality rate of this group was significantly higher than other COVID-19 hospitalized cases. Furthermore, they meaningfully experienced a prolonged stay in the hospital (p = .00). Lymphocyte count less than 20% and chest CT scan findings with more than 50% involvement might be related to the patient's mortality. These could act as laboratory and clinical indicators of disease severity, mortality, and outcome.

Apoptosis and immunophenotyping of peripheral blood lymphocytes in Iranian COVID-19 patients: Clinical and laboratory characteristics.

A novel member of human coronavirus, named severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), has been recently recognized in China and rapidly spread worldwide. Studies showed the decreasing of peripheral blood lymphocytes in a majority of patients. In this study, we have reported the clinical features, laboratory characteristics, the frequency of peripheral blood lymphocyte subpopulations, and their apoptosis pattern in Iranian coronavirus infectious disease (COVID-19) patients. Demographic and clinical data of 61 hospitalized confirmed cases with COVID-19 at Imam Khomeini Hospital were collected and analyzed. Peripheral blood mononuclear cells were isolated from all samples and the apoptosis pattern was evaluated using Annexin V/propidium iodide method. The frequency of lymphocyte subsets, including T-CD4+ , T-CD8+ , NK, B cells, and monocytes, was measured in all patients and 31 controls by flow cytometry. Our findings demonstrated that the percentage of lymphocytes, CD4+ , and CD8+ T cells were decreased in COVID-19 patients compared with the control group. Regarding the clinical severity, the number of lymphocytes, CD4+ , CD8+ T cells, and NK cells were also decreased in severe cases when compared with mild cases. Finally, our data have also indicated the increase in apoptosis of mononuclear cells from COVID-19 patients which was more remarkable in severe clinical cases. The frequency of immune cells is a useful indicator for prediction of severity and prognosis of COVID-19 patients. These results could help to explain the immunopathogenesis of SARS-CoV-2 and introducing novel biomarkers, therapeutic strategies, and vaccine candidates.

Association between pre-chemotherapy serum levels of vitamin D and clinicopathologic findings in gastric cancer.

BACKGROUND: To examine the serum levels of vitamin D in newly diagnosed gastric cancer (GC) patients compared with normal subjects and any possible association with prognostic variables. METHODS: One-hundred subjects (50 GC and 50 controls) were enrolled and serum vitamin D levels were assessed using ELISA. Based on two definitions, vitamin D was classified as a sufficient level (≥30 ng/dL) and optimal level (25-80 ng/dL). The χ2and unpaired t-test was used for data analysis with a significance level of 0.05. RESULTS: The mean serum levels of vitamin D in patients and controls were 26.86 (±14.6) and 31.72 (±13.4), respectively (P=0.09). The prevalence of vitamin D insufficiency and deficiency was higher in GC cases than controls (P=0.045 if sufficient level ≥30 and P=0.065 if sufficient level ≥25). According to histological grade analysis, grade 3 patients (poorly differentiated) were found with significantly lower vitamin D concentrations in serum than grade 1 and 2 subjects (22.25 vs 33.29 ng/dL, P=0.021). No significant differences were observed between the two groups in pathological tumor-node-metastasis (pTNM) stages, distant metastasis, and location of the tumor. CONCLUSION: Higher prevalence of vitamin D insufficiency and deficiency in GC patients may reflect its role in malignancy; however, further studies are needed to confirm this relationship and any possible benefits to the patients.

Expression Analysis of Fyn and Bat3 Signal Transduction Molecules in Patients with Chronic Lymphocytic Leukemia.

BACKGROUND: Chronic lymphocytic leukemia (CLL) is correlated with defects in T-cell function resulting imparity in antitumor immune responses. Tim-3 is a co-inhibitory immune checkpoint receptor expressed on exhausted T-cells during tumor progression. Fyn and Bat3 are two important adaptor molecules involved in inhibition and activation of Tim-3 downstream signaling, respectively. In this study, the expression of Tim-3, Fyn, and Bat3 mRNA was evaluated in CLL patients. METHODS: Peripheral blood mononuclear cells (PBMCs) were isolated from 54 patients with CLL and 34 healthy controls. Total RNA was extracted from all samples and applied for cDNA synthesis. The relative expression of Tim-3, Fyn, and Bat3 mRNA was determined by TaqMan Real-Time PCR using GAPDH as an internal control. RESULTS: Tim-3 mRNA expression was not significantly different between CLL patients and healthy controls. Fyn mRNA expression was significantly lower in CLL patients and conversely, Bat3 mRNA expression was higher in CLL patients compared to healthy controls. Interestingly, the mRNA expression of Fyn inhibitory adaptor molecule was remarkably associated with expression of Tim-3 in CLL patients. CONCLUSION: We have highlighted for the first time the expression of Fyn and Bat3 adaptor molecules in CLL patients. Our data demonstrated the strong correlation between the expression of Tim-3 and Fyn inhibitory molecules in CLL implying an important role for Tim-3-Fyn cooperation in induction of T-cell exhaustion.

A review of the immunopathogenesis of Brucellosis.

Brucellosis, caused by the intracellular pathogens Brucella, is one of the major zoonotic infections. Considering the economic burden, its prevalence has been a health concern especially in endemic regions. Brucella is able to survive and replicate within host cells by expressing different virulence factors and using various strategies to avoid the host's immune response. This leads to progression of the disease from an acute phase to chronic brucellosis. Exploration of genetic variations has confirmed the expected influence of gene polymorphisms on susceptibility and resistance to brucellosis of humans. Since there is no approved human vaccine and treatment is uncertain with risk of relapse, it is important to increase knowledge about pathogenesis, diagnosis and treatment of brucellosis in order to manage and control this infection, especially in endemic regions.

CYP2D6*3 (A2549del), *4 (G1846A), *10 (C100T) and *17 (C1023T) genetic polymorphisms in Iranian breast cancer patients treated with adjuvant tamoxifen.

There is controversy regarding the efficacy of tamoxifen in breast cancer patients who are carriers of cytochrome P450 2D6 (CYP2D6) gene polymorphisms. Poor metabolizer genotypes may not fully convert tamoxifen to its active metabolite endoxifen and thus have less exposure to anti-estrogen therapy. The present study was conducted to identify the prevalence of CYP2D6 genotypes among Iranian breast cancer patients. A total of 84 estrogen receptor-positive breast cancer patients treated at a referral center in the north of Iran were examined. A peripheral blood sample was obtained from each patient to determine the presence of *3, *4, *10 and *17 single nucleotide polymorphisms of the CYP2D6 gene by polymerase chain reaction-based restriction fragment-length polymorphism analysis. Of the four genotypes assessed, CYP2D6*4 was the most common variant and was identified in 41 (48.8%) patients as heterozygous (G/A) and 3 (3.6%) as homozygous (A/A) alleles. CYP2D6*10 heterozygous mutated alleles (C/T) were also a common genotype that presented in 22 (26.2%) of the study subjects. Variant *17 was less common and was detected only as heterozygous (C/T) in 3 patients (3.6%). No CYP2D6*3 heterozygous or homozygous mutated alleles were observed. In conclusion, the frequency of the CYP2D6 nonfunctional alleles *4 and *10 appeared relatively high in Iranian patients with hormone-sensitive breast cancer. This finding may affect the selection of an optimal hormone therapy, as patients with low CYP2D6 pathway activity may not sufficiently convert tamoxifen to its active metabolite endoxifen.

TP53 single nucleotide polymorphism (rs1042522) in Iranian patients with coronary artery disease.

Chronic diseases including coronary artery disease (CAD) impose a high burden in terms of mortality and disability particularly in developing countries. Both genetic and environmental risk factors confer susceptibility to CAD. Meanwhile, a functional polymorphism in the tumor protein p53 (TP53) gene (codon 72, exon 4) has been reported to be associated with a wide range of cancers and inflammatory disorders. There are controversies regarding CAD and involvement of the TP53 codon 72 single nucleotide polymorphism; therefore, the present case-control study was conducted to evaluate the potential association between this TP53 polymorphism and CAD in an Iranian population. A total of 153 subjects (including 70 patients diagnosed with CAD and 83 subjects with normal coronary parameters, determined by angiography) were genotyped for the TP53 (rs1042522) polymorphism by the polymerase chain reaction-restriction fragment length polymorphism technique. Clinical and laboratory findings were also evaluated. The χ2 test and unpaired Student's t-test were applied to compare genotype and allele distributions and clinical characteristics between the two groups. Significant associations of the Pro72 allele [odds ratio (OR)=1.66, P=0.027] and Pro/Pro genotype (OR=2.91, P=0.022) with CAD were identified. No associations between patients' clinical findings and genotypes were apparent. Therefore, according to present findings, the TP53 Pro72 allele may be involved in the development of CAD along with conventional risk factors in patients from Northern Iran.

Association of P53 gene polymorphism with gastric cancer in Northern Iran as a high-risk region.

Gastric cancer has the fourth highest morbidity rate of all cancers worldwide. Genetic factors including alterations in oncogenes and tumor suppressor genes serve an important role in gastric cancer development and progression. The P53 gene acts as a tumor suppressor gene by regulating the cell cycle, DNA transcription and repair, apoptosis, senescence and genome stability. In addition to somatic P53 mutations in cancer development, germline polymorphisms are also involved in different malignancies. The polymorphism of P53 at codon 72 (Arg72Pro) is established as a common variant that increases susceptibility to various cancers. The present case-control study was conducted to evaluate the possible association between this P53 polymorphism and gastric cancer in the Iranian population. A total of 59 patients with gastric cancer and 59 healthy controls were enrolled in the present study. Genomic DNA was extracted from peripheral blood mononuclear cells and genotype analysis was performed using a polymerase chain reaction-based restriction fragment length polymorphism assay. Genotype frequencies did not differ significantly between the patients and controls (P=0.4); the frequencies of the three genotypes Arg/Arg, Arg/Pro and Pro/Pro in gastric cancer patients were 28.8, 49.2 and 22.0%, and in controls were 37.3, 49.2 and 13.6%. Additionally, there were no differences in genotype frequencies based on tumor location, histological differentiation or tumor stage. Based on these findings, it may be concluded that the P53 codon 72 polymorphism does not contribute to gastric cancer susceptibility in Northern Iran.

A systematic review and meta-analysis of seroprevalence of varicella zoster virus: A nationwide population-based study.

Varicella zoster virus (VZV) causes chicken pox as a primary infection following which it becomes latent in neurons. It may then reactivate to cause shingles (herpes zoster). Severity of lesions and VZV pathogenicity are depended on the host's immune response and variant in VZV Dr Athina Myrto ChioniIdentification of VZV seroprevalance rate in general population may lead to develop new health strategic managements such as vaccination. Therefore, we aimed to provide a systematic review of the seroprevalence of VZV infection among Iranian population and estimate age- and gender- specific prevalence of VZV. Keywords "seroprevalence"; "varicella zoster virus" and "Iran"; were searched in international electronic databases and also in national Persian databases. Twenty two pooled studies among 262 total studies containing (240 published articles; 18 dissertations; and 4 proceedings abstracts) from 1992 to 2014 with total sample size of 7867 individuals were included in the final review. Data was analyzed using random effect method. The heterogeneity was calculated using I-square statistics The overall IgG seroprevalence rate of VZV infection in general population of Iran was 78.50% (95% CI; 77.74%-79.25%). There was significant heterogeneity among the studies (P<0.0001; I2=99.4%). Furthermore, the relative risk of VZV infection is high in females (80.47%, 95% CI; 79.40%-81.54%) and older adults (95.30%, 95% CI; 94.11% -96.48%). Our results may represent a true background and estimation of VZV infection in Iran and generate the cost-benefits immunization program. Moreover, the ensuing data suggests further attention on disease seroprevalence in order to obtain efficient data for therapeutic intervention targeted against VZV.

Serum Levels of Soluble CD26, A Novel Prognostic Marker for Hepatitis E Infection.

BACKGROUND: Even without treatment, most acute hepatitis E virus (HEV) infected patients resolve HEV but sometimes the disease leads to acute liver failure, chronic infection, or extrahepatic symptoms. The mechanisms of HEV pathogenesis appear to be substantially immune mediated. However, the immune responses to HEV are not precisely identified. OBJECTIVES: This study aimed to evaluate the Th1/Th2 ratio by investigating serum soluble markers from Th1 and Th2 cells in acute HEV infected patients. PATIENTS AND METHODS: This case-control study included 35 acute HEV infected patients and 35 age and gender matched anti-HEV negative healthy controls. The serum levels of Interferon (IFN)-γ, IL-4, soluble CD26 (sCD26) and sCD30 were determined by the enzyme-linked immunosorbent assay. RESULTS: The results showed a significant difference in IFN-γ and sCD26 (P < 0.0001 and P = 0.001) yet not IL-4 and sCD30 (P = 0.354 and P = 0.159) between acute HEV patients and controls, respectively. There was a positive direct correlation between serum levels of sCD26 and IFN-γ in acute HEV patients (r = 0.64, P = 0.001). In addition, the ratio of sCD26/sCD30 in the acute HEV group was more than two folds higher than in the HEV negative controls. CONCLUSIONS: Acute HEV infection shows a pattern of Th1-type immune response, and the direct significant positive correlation between the serum level of sCD26 and IFN-γ in acute HEV infected patients, suggests that the trend of sCD26 levels is a valuable marker for predicting hepatic inflammation in hepatitis E.

Data supporting the design and evaluation of a universal primer pair for pseudogene-free amplification of HPRT1 in real-time PCR.

Hypoxanthine-guanine phosphoribosyltransferase 1 (HPRT1) is a common housekeeping gene for sample normalization in the quantitative reverse transcriptase polymerase chain (qRT-PCR). However, co-amplification of HPRT1 pseudogenes may affect accurate results obtained in qRT-PCR. We designed a primer pair (HPSF) for pseudogene-free amplification of HPRT1 in qRT-PCR [1]. We showed specific amplification of HPRT1 mRNA in some common laboratory cell lines, including HeLa, NIH/3T3, CHO, BHK, COS-7 and VERO. This article provides data supporting the presence and location of HPRT1 pseudogenes within human and mouse genome, and the strategies used for designing primers that avoid the co-amplification of contaminating pseudogenes in qRT-PCR. In silico analysis of human genome showed three homologous sequences for HPRT1 on chromosomes 4, 5 and 11. The mRNA sequence of HPRT1 was aligned with the pseudogenes, and the primers were designed toward 5' end of HPRT1 mRNA that was only specific to HPRT1 mRNA not to the pseudogenes. The standard curve plot generated by HPSF primers showed the correlation coefficient of 0.999 and the reaction efficiency of 99.5%. Our findings suggest that HPSF primers can be recommended as a candidate primer pair for accurate and reproducible qRT-PCR assays.

Pseudogene-free amplification of HPRT1 in quantitative reverse transcriptase polymerase chain reaction.

Quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) provides a powerful tool for precise gene expression analysis. The accuracy of the results highly depends on careful selection of a reference gene for data normalization. HPRT1 (hypoxanthine phosphoribosyl transferase 1) is a frequently used housekeeping gene for normalizing relative expression values. However, the existence of processed pseudogenes for HPRT1 might interfere with reliable results obtained in qRT-PCR due to amplification of unintended products. Here, we designed a primer pair for pseudogene-free amplification of HPRT1 in qRT-PCR. We demonstrate that this primer pair specifically amplified HPRT1 messenger RNA (mRNA) sequence while avoiding coamplification of the pseudogenes.