Comparison of SPAG11A gene expression in infertile men with grade 1 and 2 varicocele before and after treatment.

OBJECTIVE: Sperm Associated Antigen 11A (SPAG11A) protein is a family of the epididymis-specific secretory proteins implicated in sperm maturation and function. Varicocele might cause pathophysiological difficulties in the testis and epididymis, with a harmful effect on the environment for spermatogenesis and sperm maturation. The aim of this study was to evaluate the expression level of the SPAG11A gene and sperm parameters in infertile men with grade 1 and 2 varicocele before and after treatment. METHODS: Semen specimens were collected from 20 infertile men with varicocele pre-and post-treatment and 10 healthy volunteers. Semen analysis was conducted according to world health organization guidelines. Real time PCR (qRT-PCR) reaction was applied for determination of SPAG11A mRNA expression. RESULTS: The results showed that there was a significant difference between the concentration and normal morphology between pre- and post-treatment groups and the controls. There were significant differences between pre-treatment and control groups in terms of progressive and non-progressive mobility. SPAG11A mRNA levels were significantly lower in the pre-treatment group than in healthy control subjects (p=0.007). There was no statistically significant difference in the expression of SPAG11A as well as semen parameters in the post-treatment group compared to the pre-treatment group. CONCLUSIONS: SPAG11A gene expression and semen parameters may be affected by varicocele. Whether varicocele treatment is an effective approach to reduce the adverse effect of this disease on SPAG11A expression and semen parameters needs further investigation.

The association of RBX1 and BAMBI gene expression with oocyte maturation in PCOS women.

BACKGROUND: Polycystic ovarian syndrome (PCOS) is a common endocrine disorder that affects 6-20% of women of reproductive age. One of the symptoms of PCOS is hyperandrogenism, which can impair follicular development. This disruption can cause issues with the development of oocytes and the growth of embryos. Although the exact cause of PCOS is not yet fully understood, studying the gene expression pattern of cumulus cells, which play a crucial role in the maturation and quality of oocytes, could help identify the genes associated with oocyte maturation in PCOS women. Through indirect activation of APC/Cdc20, RBX1 enables oocytes to bypass the GV (germinal vesicles) stage and advance to the MII (metaphase II) stage. our other gene is the BAMBI gene which stimulates WNT signaling, that is a crucial pathway for healthy ovarian function. This study aims to explore the expression level of the RBX1 and BAMBI genes between GV and MII oocytes of PCOS and non-PCOS groups. METHODS: In this experiment, we gathered the cumulus cells of MII (38 cases and 33 control) and GV (38 cases and 33 control) oocytes from women with/without PCOS. Besides, quantitative RT-PCR was used to assess the semi-quantitative expression of BAMBI and RBX1. RESULTS: According to our research, the expression level of RBX1 and BAMBI in MII and GV cumulus cells of PCOS patients was significantly lower than that in non-PCOS ones. CONCLUSION: This research raises the possibility of RBX1 and BAMBI involvement in oocyte quality in PCOS women.

Detrimental effects of radiofrequency electromagnetic waves emitted by mobile phones on morphokinetics, oxidative stress, and apoptosis in mouse preimplantation embryos.

Due to the increasing use of smart mobile phones, the impact of radiofrequency electromagnetic radiation (RF-EMR) on reproductive health has become a serious concern. This study investigated the effect of mobile phone RF-EMR with frequency 900-1800 MHZ on the mouse embryo morphokinetics and genotoxic effect in laboratory conditions. After ovarian stimulation in mice, the MII oocytes were collected and underwent by in vitro fertilization (IVF) method. The generated zygotes were divided into control and exposed groups. Then, the zygotes with 30 min of exposure to mobile phone RF-EMR, and the control zygotes without exposure, were incubated in the time-lapse for 5 days. The intracellular reactive oxygen species (ROS) level, morphokinetic, embryo viability rate, and Gene expression were evaluated. Exposure of zygotes to RF-EMR by inducing ROS caused a significant decrease in blastocyst viability (87.85 ± 2.86 versus 94.23 ± 2.44), delay in cleavage development (t3-t12) and also increased the time (in hours) to reach the blastocyst stage (97.44 ± 5.21 versus 92.56 ± 6.7) compared to the control group. A significant increase observed in mRNA levels of Hsp70 in exposed animals; while Sod gene expression showed a significant down-regulation in this group compared to the controls, respectively. However, there was no significant change in the transcript level of proapoptotic and antiapoptotic genes in embryos of the exposed group compared to the controls. RF-EMR emitted by mobile phone with a frequency of 900-1800 MHZ, through inducing the production of ROS and oxidative stress, could negatively affect the growth and development as well as the transcript levels of oxidative stress associated genes in the preimplantation embryos of mice.

Study Protocol for the Interactions between Dietary Patterns and ARL15 and ADIPOQ Genes Polymorphisms on Cardiometabolic Risk Factors.

Background: Cardiovascular diseases (CVDs) are recognized as one of the leading causes of death worldwide. Studies have shown the impact of genetic predisposition and dietary factors on developing these diseases. Dietary patterns and genetic factors such as polymorphisms related to the level of adiponectin may also interact with each other and produce variances in the effects of these factors on different individuals. The purpose of this study is to investigate the interactions between food intake patterns and polymorphisms on ADIPOQ and ARL15 genes in relation to cardiometabolic risk factors. Methods: This cross-sectional study is conducted on 380 adults (20 to 70 years old) living in Yazd, Iran. Individuals were selected from the participants in Yazd Health Study (YaHS) and its sub-study called Taghziyeh Mardom-e Yazd (TAMYZ) after reviewing the inclusion and exclusion criteria. YaHS is a population-based cohort study which has been conducted on 9962 adults living in Yazd since 2014. In the present study, rotated principle component analysis (PCA) with Varimax rotation is used to identify the major dietary patterns. The polymerase chain reaction-restricted fragment length polymorphism (PCR-RFLP) method is used in order to identify rs1501299 and rs6450176 variants (on ADIPOQ and ARL15 genes, respectively). General linear models (GLM) as well as regression models are used to investigate the interactions between the studied genotypes and the extracted dietary patterns. Conclusions: The results of this study can help to personalize dietary recommendations for the prevention of CVDs according to the genetic predisposition of individuals.

Misexpression of LINC01410, FOSL1, and MAFB in peripheral blood mononuclear cells associated with diabetic nephropathy.

AIMS: Currently, diabetic nephropathy (DN) is considered the leading cause of the end-stage renal disease (ESRD). However, its specific molecular mechanism is still unclear, and there is still a lack of effective diagnostic and therapeutic methods. METHOD: A pathway was assumed after bioinformatics analysis of GEO datasets related to individuals with various levels of DN, LINC01410, MAFB, and FOSL1. Then, 46 patients with type 2 diabetes (T2DM) and different levels of albuminuria, and 12 individuals without diabetes, were selected. qPCR was performed to evaluate gene expression. One-way ANOVA followed by Tukey's -and linear trend tests were performed to analyze gene expression in different stages of the disease. Moreover, receiver operating characteristic (ROC) curves and the correlation between LINC01410, FOSL1, and MAFB were analyzed. RESULTS: LINC01410, MAFB, and FOSL1 were selected based on bioinformatics analyses. The qPCR results showed that the expression of LINC01410 decreased, and FOSL1 and MAFB increased in micro-and macroalbuminuria groups compared to normoalbuminuria groups (P < 0.05). ROC curves demonstrated a significant diagnostic accuracy of LINC01410, MAFB, and FOSL1 between DN and participants with normoalbuminuria (P < 0.05). Pearson's correlation analysis revealed a positive association between the expressions of FOSL1 and MAFB (p = 0.01, r = 0.39). However, there was no correlation between LINC01410 with MAFB and FOSL1 (p = 0.23 and p = 0.21, respectively). CONCLUSION: Dysregulation of LINC01410, MAFB, and FOSL1 is related to DN. These results may provide new insights into the role of LINC01410, MAFB, and FOSL1 as potential biomarkers in DN.

Evaluation of the relationship between miR-1271 and GRB2 gene in endometriosis.

BACKGROUND: Endometriosis is a common gynecological condition with a substantial economic burden on society. It is known that both genetic and environmental factors are contributing to the phenotypic development of the disease. MicroRNAs have a vital role in the pathogenesis of endometriosis. miR-1271 and its direct target gene, GRB2 (growth factor receptor-bound protein 2), expression have been studied in gynecologic cancers, while their role in endometriosis has not been studied. OBJECTIVE: We measured miR-1271 and GRB2 gene expression in the eutopic and ectopic tissues of patients (endometrial tissues) in contrast to the control samples from healthy women. MATERIALS AND METHODS: In this study, a total of 45 samples (15 control samples, 15 eutopic samples and 15 ectopic samples) were collected. We used qRT-PCR (quantitative polymerase chain reaction) to evaluate the expression levels of the miR-1271 and GRB2 gene. RESULTS: We observed inverse expression of miR-1271 and GRB2 gene. MiR-1271 expression was significantly reduced in patients with endometriosis compared with healthy women. While there was a noticeable increase in the expression level of its target gene, GRB2, in tissues of endometriosis patients compared with normal control samples. CONCLUSION: We have shown an inverse relationship between the reduction of miR-1271 expression level and increase in the expression level of GRB2, therefore, increased GRB2 expression in endometriosis tissues can be due to decreased expression of this microRNA. Our findings suggested that miR-1271 maybe play a role as a biomarker in the diagnosis of patients with endometriosis.

Novel variants underlying autosomal recessive neurodevelopmental disorders with intellectual disability in Iranian consanguineous families.

BACKGROUND: Intellectual disability (ID) is a heterogeneous group of neurodevelopmental disorders that is characterized by significant impairment in intellectual and adaptive functioning with onset during the developmental period. Whole-exome sequencing (WES)-based studies in the consanguineous families with individuals affected with ID have shown a high burden of relevant variants. So far, over 700 genes have been reported in syndromic and non-syndromic ID. However, genetic causes in more than 50% of ID patients still remain unclear. METHODS: Whole-exome sequencing was applied for investigation of various variants of ID, then Sanger sequencing and in silico analysis in ten patients from five Iranian consanguineous families diagnosed with autosomal recessive neurodevelopmental disorders, intellectual disability, performed for confirming the causative mutation within the probands. The most patients presented moderate-to-severe intellectual disability, developmental delay, seizure, speech problem, high level of lactate, and onset before 10 years. RESULTS: Filtering the data identified by WES, two novel homozygous missense variants in FBXO31 and TIMM50 genes and one previously reported mutation in the CEP290 gene in the probands were found. Sanger sequencing confirmed the homozygote variant's presence of TIMM50 and FBXO31 genes in six patients and two affected siblings in their respective families. Our computational results predicted that the variants are located in the conserved regions across different species and have the impacts on the protein stability. CONCLUSION: Hence, we provide evidence for the pathogenicity of two novel variants in the patients which will expand our knowledge about potential mutation involved in the heterogeneous disease.

Plasma miR-21 as a potential predictor in prediabetic individuals with a positive family history of type 2 diabetes mellitus.

Type 2 diabetes mellitus (T2DM) is a heritable metabolic perturbation, rapidly growing across the world. Primary recognition of susceptible individuals with a family history of type 2 diabetes (FHD) in the prediabetes stage could delay the onset of T2DM or reduce complications induced by diabetes. This study aims to evaluate the expression levels of miR-21, miR-126 as noninvasive predictive biomarkers in individuals with genetic predisposition and investigate the correlation of miRNAs and cardiometabolic risk factors. Our study demonstrated that miR-21 expression has a notable elevate in both groups of T2DM and pre-T2DM. miR-21 expression was distinguished in the pre-T2DM and T2DM from the nondiabetic individuals by ROC curve analysis with AUC of 0.77 (95% CI 0.65-0.90; p = 0.0004) and AUC of 0.78 (95% CI 0.64-0.92; p = 0.0042), respectively. The relative gene expression of miR-126 was nearly equal among groups. miR-21 expression was positively associated with glycosylated hemoglobin (HbA1c), fasting blood sugar (FBS), and triglyceride (TG) and might have diagnostic value for T2DM and pre-T2DM. This study has revealed that the expression level of miR-21 can be considered as a non-invasive and rapid tool for distinguishing pre-T2DM and T2DM counterparts from healthy individuals.

Appropriate fixative for MEM-G/9 staining of cultured human HLA-G-positive JEG-3 trophoblast tumor cells.

Human leukocyte antigen (HLA-G) participates in immunosuppression and is useful for prenatal diagnostics. Isolation of fetal cells positive for HLA-G by HLA-G antibody conjugated nanoparticles from the cervix of pregnant women is the basis for non-invasive prenatal testing. Endocervical specimens are fixed in transport medium before isolation using antibody conjugated nanoparticles. Staining of HLA-G using MEM-G/9 antibody, however, is restricted to unfixed cells. We investigated the effect of several fixatives on the interaction of HLA-G with MEM-G/9 in the HLA-G-positive cell line, JEG-3. We investigated absolute methanol, 1:1 acetate buffer:methanol, Pap solution and paraformaldehyde. The effects of these fixatives were evaluated using immunofluorescence. We found no MEM-G/9 surface staining of methanol fixed cells. Approximately 40% of JEG-3 cells fixed with paraformaldehyde failed to stain. Nearly all cells were stained with MEM-G/9 following fixation with acetate buffer:methanol or Pap solution. Our findings indicate the importance of using an appropriate fixative for preserving HLA-G cell surface antigen for studies using the MEM-G/9 antibody.

CAG repeat polymorphism in androgen receptor and infertility: A case-control study.

BACKGROUND: Androgens play a role in the development of male phenotype and spermatogenesis during puberty, the function of which is regulated by the androgen receptor (AR) gene. There is a polymorphism site in exon 1 of the gene encoding this receptor that can have different frequencies of CAG trinucleotide repeats and leads to the formation of polyglutamine chains of different lengths in the N-terminal domain of the AR protein and reduced sperm production by affecting spermatogenesis. OBJECTIVE: To investigate whether the cause of a group of unexplained infertilities could be the increased frequency of CAG repeats in the AR gene of patients with oligozoospermia and azoospermia. MATERIALS AND METHODS: In this case-control study, 84 men including 42 with unexplained infertility As a case group and 42 fertile men as a control group were selected. The frequency of CAG repeats was determined by the polymerase chain reaction method and then the difference in the frequency of these repeats was determined based on the difference in band size on the agarose gel. RESULTS: The mean CAG repeat length in the azoospermia and oligozoospermia group was 17.5 ± 0.63 and in the fertile group it was 16.11 ± 0.75 (p = 0.46). In addition, most men (88.1% in the case group and 71.41% in the control group) had 13-23 repeats. CONCLUSION: No significant correlation was found between CAG repeat length and the risk of male factor infertility in an ethnically defined population of Iranian men. The role of regulatory factors and epigenetic changes should be taken into account too.

Circulating microRNA-122, microRNA-126-3p and microRNA-146a are associated with inflammation in patients with pre-diabetes and type 2 diabetes mellitus: A case control study.

The prevalence of type 2 diabetes mellitus (T2DM) is increasing dramatically worldwide. Dysregulation of microRNA (miRNA) as key regulators of gene expression, has been reported in numerous diseases including diabetes. The aim of this study was to investigate the expression levels of miRNA-122, miRNA-126-3p and miRNA-146a in diabetic and pre-diabetic patients and in healthy individuals, and to determine whether the changes in the level of these miRNAs are reliable biomarkers in diagnosis, prognosis, and pathogenesis of T2DM. Additionally, we examined the relationship between miRNA levels and plasma concentrations of inflammatory factors including tumor necrosis factor alpha (TNF-α) and interleukin 6 (Il-6) as well as insulin resistance. In this case-control study, participants (n = 90) were allocated to three groups (n = 30/group): T2DM, pre-diabetes and healthy individuals as control (males and females, age: 25-65, body mass index: 25-35). Expression of miRNA was determined by real-time polymerase chain reaction (RT-PCR). Furthermore, plasma concentrations of TNF-α, IL-6 and fasting insulin were measured by enzyme-linked immunosorbent assay. Homeostatic model assessment for insulin resistance (HOMA-IR) was calculated as an indicator of insulin resistance. MiRNA-122 levels were higher while miRNA-126-3p and miRNA-146a levels were lower in T2DM and pre-diabetic patients compared to control (p<0.05). Furthermore, a positive correlation was found between miRNA-122 expression and TNF-α (r = 0.82), IL-6 (r = 0.83) and insulin resistance (r = 0.8). Conversely, negative correlations were observed between miRNA-126-3p and miRNA-146a levels and TNF-α (r = -0.7 and r = -0.82 respectively), IL-6 (r = -0.65 and r = -0.78 respectively) as well as insulin resistance (r = -0.67 and r = -0.78 respectively) (all p<0.05). Findings of this study suggest the miRNAs can potentially contribute to the pathogenesis of T2DM. Further studies are required to examine the reproducibility of these findings.

Atezolizumab and granzyme B as immunotoxin against PD-L1 antigen; an insilico study.

CD274 gene encodes programmed death-ligand 1 (PD-L1) protein, also known as B7 homolog 1 (B7-H1), which is a crucial hallmark for highly proliferation cells including cancer cells. PD-1 and PD-L1 interaction is assumed as a negative regulator for immune response which can inhibit the T cell growth and cytokine secretion and supports tumor cells evasion from immune system. therefore, PD-L1 could be assumed as a candidate target for immune-therapy. The predicted structure of PD-L1 indicates (Gly4Ser) 3 linker-based chains links. In that line, different simulation softwares applied to explore the structure of granzyme B (GrB), a serine protease in cytotoxic lymphocytes granules as an apoptosis mediator, was attached to its specific antibody structure (atezolizumab) via an adaptor sequence. Evaluation of accuracy, energy minimization and characterization of biological properties of the final processed structure were performed and our computational outcomes indicated that the employed method for structure prediction has been successfully managed to design the immunotoxin structure. It is necessary to mention that, the precise and accurate design of the immune-therapeutic agents against cancer cells can be confirmed by employment of in-silico approaches. Consequently, based on this approach we could introduce a capable immunotoxin which specifically targeting PD-L1 in an accurate orientation and initiates cancer cell destruction by its toxin domain. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s40203-021-00076-z.

Co-delivery of miRNA-15a and miRNA-16-1 using cationic PEGylated niosomes downregulates Bcl-2 and induces apoptosis in prostate cancer cells.

OBJECTIVE: Tumor suppressor miRNAs, miR-15a and miR-16-1, with high-specificity and oncogenic targeting of Bcl-2, can target tumor tissues. Disadvantages of the clinical application of free miRNAs include poor cellular uptake and instability in plasma, which can be partially improved by using nanocarriers to deliver anti-cancer agents to the tumor cell. METHOD: In this study, cationic niosomes were designed and optimized with the specific formulation. Then, the physical characteristics, the cytotoxicity, the impact of transfected miRNAs on the expression of the Bcl-2 gene, and the apoptosis rate of the different formulation into prostate cancer cell were determined. RESULTS: The optimum formulation containing tween-60: cholesterol: DOTAP: DSPE-PEG2000 at 70:30:25:5 demonstrated that the vesicle size and zeta potentials were 69.7 nm and + 14.83 mV, respectively. Additionally, noisome-loaded miRNAs had higher toxicity against cancer cells comparing with free forms. The transfection of PC3 cells with the combination therapy of nanocarriers loaded of two miRNAs led to a significant decrease in the expression of the Bcl-2 gene and increased the degree of cell death in PC3 cells compared with other treatment groups, and the synergistic effects of co-delivery of miR-15a and miR-16-1 on prostate cancer cells were shown. CONCLUSION: According to the results, it seems the designed niosomes containing miR-15a and miR-16-1 can target the Bcl-2 gene and provide a cheap, applicable, cost-effective, and safe drug delivery system against prostate cancer.

Isolation of HLA-G+ cells using MEM-G/9 antibody-conjugated magnetic nanoparticles for prenatal screening: a reliable, fast and efficient method.

The development of an effective and noninvasive early method for obtaining fetal cells is crucial to prenatal screening. Despite proving the presence of fetal cells in the reproductive tract, their use is limited due to their inability to properly isolate them from maternal cells. Magnetic-activated cell sorting (MACS) is a simple technique to separate cells. The present study aimed to develop a MACS-based platform for the isolation of the HLA-G expressing trophoblast cells. For this purpose, first, the triazine functionalized MNPs were synthesized and characterized. Then, MNPs were directly and indirectly conjugated by the MEM-G/9 antibodies targeting HLA-G+ cells. The antibody amount on the surface of the nanoparticles was determined with the Bradford assay. The cell capture efficiency was also investigated. Various characterization methods confirmed the successful nanoparticle synthesis and antibody conjugation. The optimal initial antibody amount for the immobilization was about 20 µg and the optimal time was 3 h. The antibody-nanoparticles by the indirect method had better targeting and capture efficiency than the direct method. The MNPs indirectly conjugated with antibodies are an efficient tool for cell isolation and present considerable potential to be applied in biomedical fields.

Unraveling the dark matter, long non-coding RNAs, in male reproductive diseases: A narrative review.

Recent advances in human transcriptome have revealed the fundamental and functional roles of long non-coding RNA in the susceptibility to diverse diseases and pathological conditions. They participate in wide range of biological processes such as the modulating of chromatin structure, transcription, translation, and post-translation modification. In addition, based on their unique expression profiles and their association with clinical abnormalities such as those of related to male reproductive diseases, they can be used to develop therapeutic methods and biomarkers for screening of the diseases. In this study, we will review the identified lncRNAs and their molecular functions in the pathogenesis of male reproductive diseases such as prostate cancer, benign prostatic hyperplasia, prostatitis, testicular cancer, varicocele, and sperm abnormalities.

Reanalysis of discarded blastocysts for autosomal aneuploidy after sex selection in cleavage-stage embryos.

OBJECTIVE: The goal of the present study was to investigate the rate of chromosomal aneuploidies in surplus embryos after sex determination at the cleavage stage. Then, the same chromosomal aneuploidies were evaluated in blastocysts after extended culture. METHODS: Sixty-eight surplus embryos were biopsied at the cleavage stage and incubated for an additional 3 days to allow them to reach the blastocyst stage. The embryos were reanalyzed via fluorescence in situ hybridization (FISH) to examine eight chromosomes (13, 15, 16, 18, 21, 22, X, and Y) in both cleavage-stage embryos and blastocysts. RESULTS: Although the total abnormality rate was lower in blastocysts (32.35%) than in cleavage-stage embryos (45.58%), the difference was not significant (p=0.113). However, when we restricted the analysis to autosomal abnormalities, we observed a significant difference in the abnormality rate between the cleavage-stage embryos (44.11%) and the blastocysts (17.64%, p=0.008). A higher rate of sex chromosomal abnormalities was also observed in cleavage-stage embryos (29.4%) than in blastocysts (14.70%, p=0.038). CONCLUSION: The data indicated that embryo biopsy should be conducted at the blastocyst stage rather than the cleavage stage. The results also emphasized that examination of common chromosomal aneuploidies apart from sex selection cycles can be conducted in the blastocyst stage with the FISH method.

Investigating the expressions of miRNA-125b and TP53 in endometriosis. Does it underlie cancer-like features of endometriosis? A case-control study.

BACKGROUND: Endometriosis is generally considered as a benign condition; however, there is a possibility for it to become cancerous. miR-125b is upregulated in both endometriotic tissues and serum samples of women with endometriosis but its potential targets in endometriosis are still not fully understood. OBJECTIVE: The role of miR-125b in the regulation of TP53 expression in endometriosis was tested with a bioinformatics approach. In addition, the expression of miR-125b and TP53 in both eutopic (Eu-p) and ectopic endometrium (Ec-p) in the endometrium tissues of women with endometriosis was compared to those in the normal endometrium tissues of controls (Normal). MATERIALS AND METHODS: In this case-control study, the Eu-p and Ec-p samples were collected from 20 women who underwent laparoscopic surgery, and the normal endometrium tissues were collected from 20 controls with no evidence of endometriosis. For bioinformatics approach, a protein-protein interaction network was constructed based on co-expressed potential targets of miR-125b. Quantitative polymerase chain reaction technique was used for the measurement of miR125b and TP53 expression. RESULTS: Our results showed that miR-125b was significantly overexpressed in Ec-p (p-value: 0.021). In addition, there was a significant TP53 under expression in both the Ec-p and Eu-p samples compared with the Normal tissues (p-value: 0.003). CONCLUSION: The negative correlation between miR-125b and TP53 as well as a noticeable decreased expression of TP53 in both Ec-p and Eu-p samples may be interpreted as the roles of miR-125b/TP53 axis in the pathogenesis of endometriosis. In addition, these findings and bioinformatic analyses imply a possible role of miR-125b in cancer-like features of endometriosis.

Comparison of chromosomal instability of human amniocytes in primary and long-term cultures in AmnioMAX II and DMEM media: A cross-sectional study.

BACKGROUND: The genomic stability of stem cells to be used in cell therapy and other clinical applications is absolutely critical. In this regard, the relationship between in vitro expansion and the chromosomal instability (CIN), especially in human amniotic fluid cells (hAFCs) has not yet been completely elucidated. OBJECTIVE: To investigate the CIN of hAFCs in primary and long-term cultures and two different culture mediums. MATERIALS AND METHODS: After completing prenatal genetic diagnoses (PND) using karyotype technique and chromosomal analysis, a total of 15 samples of hAFCs from 650 samples were randomly selected and cultured in two different mediums as AmnioMAX II and DMEM. Then, proliferative cells were fixed on the slide to be used in standard chromosome G-banding analysis. Also, the senescent cells were screened for aneuploidy considering 8 chromosomes by FISH technique using two probe sets including PID I (X-13-18-21) & PID II (Y-15-16-22). RESULTS: Karyotype and interphase fluorescence in situ hybridization (iFISH) results from 650 patients who were referred for prenatal genetic diagnosis showed that only 6 out of them had culture- derived CIN as polyploidy, including mosaic diploid-triploid and diploid-tetraploid. Moreover, the investigation of aneuploidies in senesced hAFCs demonstrated the rate of total chromosomal abnormalities as 4.3% and 9.9% in AmnioMAX- and DMEM-cultured hAFCs, respectively. CONCLUSION: hAFCs showed a low rate of CIN in two AmnioMAX II and DMEM mediums and also in the proliferative and senescent phases. Therefore, they could be considered as an attractive stem cell source with therapeutic potential in regenerative medicine.

A comparative analysis of immunomodulatory genes in two clonal subpopulations of CD90+ amniocytes isolated from human amniotic fluid.

OBJECT: Immunosuppressive and immunomodulatory activity of mesenchymal stem cells derived from different sources, such as placental membranes, umbilical cord, and amniotic fluid has been proved. The heterogeneous nature of human amniocytes have been confirmed due to different clonal subpopulations found in amniotic fluid. The aim of this study was to investigate a 17-gene panel of immunomodulatory markers in two clonal subpopulations of CD90+ amniocytes, divided based on morphology into epithelioid and fibroblastoid cells. METHOD: Semi-quantitative RT-PCR was used to study the expression of the chosen genes. Flow cytometry analysis confirmed the non-hematopoietic mesenchymal origin of isolated cells, based on lacking the hematopoietic marker of CD31, and the presence of mesenchymal marker of CD90 (both on more than 90% of cells). RESULTS: Our results showed that besides growth characteristics, the two cell groups were different in expressional profile, so that, fibroblastoid clones displayed higher level of immunosuppression genes as well as mesenchymal surface marker of CD90 compared to epithelioid ones. Our previous investigation on these clones showed that epithelioid cells have a more potential to express the pluripotency genes. It seems there is an inverse relationship between genes associated with immunosuppression and pluripotency. CONCLUSION: Although many reports have been published regarding the immunosuppressive properties of fetal stem cells, but few studies to date have explained whether the stemness state of human amniocytes may affect their immunosuppressive potential. Further study on amniocytes, which often has self-renewal ability and high immunomodulatory potential, can help to understand the details of this relationship.

Effects of phosalone plant pesticide on sperm parameters and sexual hormone levels in Wistar rats: An experimental study.

BACKGROUND: Phosalone is an organophosphate insecticide, applied to control of plant pests. This compound has various side effects because it acts as an acetyl cholinesterase enzyme inhibitor. OBJECTIVE: To investigate the effects of phosalone on the sperm parameters of and levels of sex hormones in adult male rats. MATERIALS AND METHODS: In this experimental study, 16 adult (8-12 wk) male Wister rates (weighing 220-280 gr) were randomly assigned into 4 groups (n = 4/each). Group 1 (control) received only routine adequate water and food; Group 2, 3, and 4 received different low doses of phosalone (60, 90, and 120 mg/kg respectively). The rats were weighed and anesthetized after 48 days. Sperm parameters including number, motility, and viability as well as sex hormones (such as Luteinizing Hormone, Follicle Stimulating Hormone, and testosterone) were evaluated and compared after removing the epididymis tail. RESULTS: Our results showed that phosalone decreased sperm motility, viability, and number in a dose-dependent manner. The level of FSH and LH was increased, and testosterone was decreased. Also, depending on the dose, phosalone decrease sperm motility and viability (p ≤ 0.001), while the level of FSH and LH was increased and testosterone was decreased (p = 0.861). CONCLUSION: Phosalone has negative effects on reproductive indices in male rats and can cause serious damage and decrease the number and sperms motility. It can also cause infertility due to changing the concentration of hormones.