The important biological roles of Syncytin-1 of human endogenous retrovirus W (HERV-W) and Syncytin-2 of HERV-FRD in the human placenta development.

BACKGROUND:  Human endogenous retroviruses (HERVs) entered the germ line by retroviral infection from a distant ancestor over 30 million years ago and constitute 8% of the human genome. The majorities of HERVs are non-protein coding and lack function because of the accumulation of mutations, insertions, deletions, and/or truncations. However, a small number of HERV genes carried ORFs with beneficial functions for the host. METHODS & RESULTS: In this review, we summarize the structural and important biological roles of two HERV gene products termed Syncytin-1 and Syncytin-2 in human placenta development. Indeed, two retroviral gene products that have important roles in mammalian development, Syncytin-1 (HERV-W) and Syncytin-2 (HERV-FRD), are prime examples encoded by env genes and expressed in the placental trophoblasts. Several pivotal studies revealed that Syncytins are fundamental genes implicated in regulating trophoblast fusion and placenta morphogenesis. CONCLUSION:  Interestingly, it has been suggested that syncytins may also be implicated in non-fusogenic activities leading to apoptosis, proliferation, and immunosuppressive activities.

Prenatally detected congenital medulloblastoma.

We describe a congenital cerebellar mass in a fetus at 30 weeks GA. The lesion is detected at the prenatal third-trimester ultrasound, confirmed by fetal MRI, and determined as medulloblastoma in postmortem pathologic evaluation.

Histological Evaluation of Products of Conception, Who Benefits from It?

Background: Histopathological evaluation of the first trimester pregnancy loss has always been controversial. Although it is recommended, it is not a part of guidelines.Methods: Six hundred eighty-six samples in a referral infertility clinic were evaluated microscopically and categorized. Two hundred ninety-five cases were evaluated by genetic methods (Multiplex Ligation-dependent Probe Amplification).Results: From 569 samples with chorionic villi, 361 cases had history of three or more abortions. 18.3% of this group showed chronic intervillous of unknown etiology (CIUE) and 8.3% revealed intervilli fibrin deposition, both pathologies with a high risk of recurrence. History of a live child was significantly higher in CIUE group. 29% of genetically evaluated cases had a chromosomal abnormality.Conclusion: Histological evaluation of recurrent pregnancy loss could illuminate the cause of abortion in relatively acceptable percentage of cases, especially in mothers with higher number of previous abortion, mothers with a history of live child and in referral centers.

A retrospective autopsy study of 42 cases of stillbirth in Avicenna Research Institute.

BACKGROUND: According to the World Health Organization about 2.6 million deaths were reported worldwide in 2015. More than 98% of stillbirths occur in developing countries. At present, the causes of many cases of stillbirth are unknown due to the lack of necessary data and autopsies in Iran. The aim of this study was to investigate the most plausible cause of stillbirth by evaluating clinical records and autopsies. METHODS: A cross-sectional study of 42 stillbirth autopsies in Avicenna Research Institute from 2012 to 2019, was conducted. Data were extracted from a checklist prepared by the project researchers. The checklist contains maternal demographic information, medical history and maternal illness, pregnancy risk factors, placenta and stillbirth information. Collected data were reviewed and classified according to the ReCoDe (Relevant Condition at Death) system. RESULTS: In the present study, based on ReCoDe classification, related causes of 95.2% of stillbirths were identified and 4.8% were in the unclassified group. The most common causes were: Fetal causes (64.3%), umbilical cord (14.3%), placenta (7.1%), amniotic fluid (4.8%), maternal medical conditions (2.4%). The causes of about 70% of stillbirth in Iran are unexplained, but in this study, using autopsy results and ReCoDe classification, only 4.8% of stillbirth causes remained unexplained. CONCLUSIONS: In our study, unknown cases were rare after autopsy. But considering the limitations and costs of autopsy, we need to design the guideline to specify cases who need an autopsy. Fetal autopsy, placental examination and clinical information could reduce the proportion of stillbirths that remain unexplained.

Molecular analysis of testis biopsy and semen pellet as complementary methods with histopathological analysis of testis in non-obstructive azoospermia.

PURPOSE: Non-obstructive azoospermia (NOA) is one the many causes of male infertility (10 %) resulting from testicular failure. Multiple testicular biopsies fail to find mature sperm in at least 50 % of cases Therefore; hunting for sensitive and specific biomarkers of spermatogenesis that could better determine the fertility status in NOA can lead to improved management of male infertility. Therefore, we evaluated sperm production through analyses of germ cell-specific transcripts (DAZ, TSPY1, SPTRX3 and SPTRX1) in semen and testicular biopsies of men with azoospermia. METHODS: We collected semen (N=83) and testis biopsies (N=31) from men with non-obstructive azoospermia. We later extracted RNA and synthesized cDNA using washed semen precipitate and testicular tissues. We also performed semi-nested PCR with designed specific primers. Using H&E method, an expert pathologist performed the histopathological evaluation. Having categorized the patients into three groups based on histopathological results, we calculated the agreement between molecular results of semen and tissues with histopathological findings for each patient using Kappa statistical test. RESULTS: Molecular findings of precipitated semen and testicular tissues were in disagreement with histopathological results in most cases. Molecular analysis of testis biopsies showed significant difference (Kappa coefficient=0.009, P value=0.894) with histopathological results; TSPY1, DAZ, SPTRX3 and SPTRX1 were respectively detected in 94 %, 94 %, 17.6 % and 52.9 % of men diagnosed with germ cell aplasia. CONCLUSIONS: Molecular analysis of semen does not provide sufficient sensitivity and specificity to be used as a screening test at the present time, but it is a useful adjunct to histopathological methods in men with NOA. Spermatid/sperm specific transcripts indicated the possibility to find mature sperm following repeated multiple testicular sperm extraction (TESE) or microdisection TESE (mTESE).

Preclinical assessment of a recombinant RBD-Fc fusion protein as SARS-CoV-2 candidate vaccine.

Background: Waning immunity and emergence of new variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), highlight the need for further research in vaccine development. Methods: A recombinant fusion protein containing the receptor-binding domain (RBD) fused to the human IgG1 Fc (RBD-Fc) was produced in CHO-K1 cells. RBD-Fc was emulsified with four adjuvants to evaluate its immunogenicity. The RBD-specific humoral and cellular immune responses were assessed by ELISA. The virus neutralizing potency of the vaccine was investigated using four neutralization methods. Safety was studied in mice and rabbits, and Antibody-Dependent Enhancement (ADE) effects were investigated by flow cytometry. Results: RBD-Fc emulsified in Alum induced a high titer of anti-RBD antibodies with remarkable efficacy in neutralizing both pseudotyped and live SARS-CoV-2 Delta variant. The neutralization potency dropped significantly in response to the Omicron variant. RBD-Fc induced both TH2 and particularly TH1 immune responses. Histopathologic examinations demonstrated no substantial pathologic changes in different organs. No changes in serum biochemical and hematologic parameters were observed. ADE effect was not observed following immunization with RBD-Fc. Conclusion: RBD-Fc elicits highly robust neutralizing antibodies and cellular immune responses, with no adverse effects. Therefore, it could be considered a promising and safe subunit vaccine against SARS-CoV-2.

The molecular footprints of BK virus in the product of conception over the second and third gestational trimesters.

OBJECTIVE: Prior studies have shown controversial results on the vertical transmission of BK virus (BKV). The present study aimed to assess the possibility of BKV vertical transmission from mother to fetus in the product of conception (embryo, fetuses, and/or placentas) over the three stages of pregnancy. RESULTS: Of the 26 placental studied tissues, 6 were in the first trimester, and none of which were positive. Only one out of the 13 (7.7%) placental materials in the second trimester was positive. Only one out of 7 (14%) placental materials of the third trimester was positive. There were cases that no virus was detected in their placental but BKV was detected in their other tissues. Among 26 conceptuses, 17 (65%) were negative for BKV and 9 (34.6%) were positive, 7/13 (54%) were positive in the second, and 2/7 (29%) were positive in the third trimester fetuses. BKV was most frequently detected in the liver (eight cases), heart (three cases), and placenta (2 cases). There were cases that no virus was detected in their placental but BKV was detected in their other tissues.

Morphometric and immunohistochemical analysis as a method to identify undifferentiated spermatogonial cells in adult subjects with Klinefelter syndrome: a cohort study.

OBJECTIVE: To study the prevalence of spermatogonia in adult subjects with Klinefelter syndrome (KS) using MAGE-A4 and UCHL1 (PGP9.5) immunohistochemistry as markers for undifferentiated spermatogonial cells. We aimed to compare this method to the gold standard of hematoxylin and eosin (H & E) staining with histologic analysis in the largest reported cohort of adult subjects with KS. DESIGN: A retrospective cohort study. SETTING: Infertility Clinic and Institute for Regenerative Medicine. PATIENT(S): This study consisted of 79 adult subjects with KS and 12 adult control subjects. INTERVENTION(S): The subjects with KS (n = 79) underwent bilateral testicular biopsy in an initial effort to recover spermatozoa for in vitro fertilization and intracytoplasmic sperm injection. The institutional review board approved the use of a portion of the archived diagnostic pathology paraffin blocks for the study. The samples were superimposed onto microscopic slides and labeled with the PGP9.5 and MAGE-A4 antibodies. Subjects (n = 12) who had previously consented to be organ donors via the National Disease Research Interchange were selected as controls. Dedicated genitourinary pathologists examined the H & E-, PGP9.5-, and MAGE-A4-stained tissue for presence of undifferentiated spermatogonia and spermatozoa with the use of a virtual microscopy software. MAIN OUTCOME MEASURE(S): The primary outcome was the presence of MAGE-A4-positive or UCHL1-positive tubules that indicate undifferentiated spermatogonia. Supportive outcomes include assessing the biopsy specimen for the following: total surface area; total seminiferous tubule surface area; total interstitium surface area; the total number of seminiferous tubules; and MAGE-A4- negative or UCHL1-negative tubules. Additionally, clinical information, such as age, karyotype, height, weight, mean testicle size, and hormonal panel (luteinizing hormone, follicle-stimulating hormone, and testosterone), was obtained and used in a single and multivariable analysis with linear regression to determine predictive factors for the number of UCHL1-positive tubules. RESULT(S): The mean age of the subjects in the KS group was 32.9 ± 0.7 years (range, 16-48). UCHL1 (PGP9.5) and MAGE-A4 staining showed that 74.7% (n = 59) and 40.5% (n = 32) of the subjects with KS, respectively, were positive for undifferentiated spermatogonia compared with 100% (n = 12) of the control subjects who were positive for both the markers. Hematoxylin and eosin with microscopic analysis showed that only 10.1% (n = 8) of the subjects were positive for spermatogonia. The mean number of positive tubules per subject with KS was 11.8 ± 1.8 for UCHL1 and 3.7 ± 1.0 for MAGE-A4. Secondary analysis showed 7 (8.9%) adult subjects with KS as positive for spermatozoa on biopsy. The population having negative testicular sperm extraction results (n = 72) showed a spermatogonia-positive rate of 1.4%, (n = 1), 72.2% (n = 52), and 34.7% (n = 25) using H & E, UCHL1, and MAGE-A4, respectively. Further analysis showed that 54 (75.0%) subjects were either positive for UCHL1 or MAGE-A4. Twenty (27.8%) subjects were positive for both UCHL1 and MAGE-A4. Multivariate analysis with linear regression showed no significant correlation between clinical variables and the number of UCHL1-positive tubules found on biopsy specimens. CONCLUSION(S): We report a cohort of adult subjects with KS undergoing analysis for the presence of undifferentiated spermatogonia. UCHL1 and MAGE-A4 immunostaining appear to be an effective way of identifying undifferentiated spermatogonia in testicular biopsy specimens of subjects with KS. Despite observing deterioration in the testicular architecture, many patients remain positive for undifferentiated spermatogonia, which could be harvested and potentially used for infertility therapy in a patient with KS who is azoospermic and has negative testicular sperm extraction results.

Detection of Aneuploidies in Products of Conception and Neonatal Deaths in Iranian Patients Using the Multiplex Ligation-Dependent Probe Amplification (MLPA).

BACKGROUND: Around 70% of all pregnancies (Including 15% of clinically-recognized ones) are lost due to various fetal or maternal disorders. Chromosomal aneuploidies are among the most common causes of pregnancy loss. Standard chromosome analysis using G-banding technique (Karyotype) is the technique of choice in studying such abnormalities; however, this technique is time-consuming and sensitive, and limited by vulnerabilities such as cell culture failure. The use of molecular cytogenetic techniques, including array-based techniques and Multiplex Ligation-Dependent Probe Amplification (MLPA), has been proposed to overcome the limitations of this method to study the products of conception. This study has been designed to investigate the feasibility of using MLPA technique as a standalone genetic testing, with histopathologic examinations and genetic counseling to detect aneuploidies in products of conception and neonatal deaths. METHODS: Forty-two verified fetal and neonatal samples were studies and genetic counseling was scheduled for all parents. Histopathologic examinations were carried out on the products of conception, and appropriate fetal tissues were separated for genetic studies. Following DNA extraction and purification, MLPA was carried out to investigate chromosomal aneuploidies. RESULTS: Nine samples (21.42%) were diagnosed to be affected with aneuploidy. Detected aneuploidies were trisomy 22 (n=3), trisomy 21(n=1), trisomy 18 (n=2), trisomy 16 (n=1), trisomy 13 (n=1), and monosomy of chromosome X (n=1). The MLPA analysis results were conclusive for all of the fetal samples (Success rate: 100%). CONCLUSION: These results suggest that MLPA, as a standalone genetic testing, is an accurate, rapid, and reliable method in overcoming the limitations of standard cytogenetic techniques in genetic investigation of products of conception.

Thyroid peroxidase in human endometrium and placenta: a potential target for anti-TPO antibodies.

Autoimmune thyroid disease is the most common endocrine disorder during pregnancy. Thyroid autoantibodies (TAs) have been suggested to serve a role in implantation failure and spontaneous abortion. Until now, there are no data on the potential interaction of TAs with human reproductive organs. Here, we set out for the first time to test this hypothesis by studying the expression of thyroid peroxidase (TPO) at gene and protein level in human reproductive organs. Endometrial samples were taken from normal women, and placenta tissues were collected after full-term caesarian section. Expression of TPO messenger RNA (mRNA) was investigated by qRT-PCR. In addition, polyclonal anti-TPO antibodies were produced and the expression of TPO protein in mentioned tissues was evaluated by immunohistochemistry and Western blot analysis. The reactivity of anti-TPO antibody in human embryos was evaluated by immunofluorescent staining. For the first time, our study showed that TPO is expressed at gene and protein levels in endometrium and placenta. TPO expression was mainly localized to glandular and luminal epithelial cells in the endometrium. In placenta, the syncytiotrophoblasts and invasive trophoblast cells were the main cell types that expressed TPO protein. Specific band of approximately 110 kDa was observed in all endometrial and placental tissues by Western blot analysis. However, no expression of TPO protein was observed in human embryo. TPO expression in endometrium and placenta may explain higher frequency of abortion and infertility in patients with thyroid autoimmunity.

Effects of silymarin, cabergoline and letrozole on rat model of endometriosis.

OBJECTIVE: Silymarin as an herbal drug has potent antioxidant effects that could make it a good choice for endometriosis therapy. The aim of the current study was to determine the effects of silymarin as an herbal drug on induced endometrial lesion in rat model of endometriosis. MATERIALS AND METHODS: A total of 32 mature, female Sprague-Dawley rats were allocated into 4 experimental groups. The duration of study was about 6 months. Endometriosis implants were surgically prepared and autografted into 32 rats. Three weeks after endometriosis induction, animals were randomly allocated into four groups: Group 1 received cabergoline (CAB group); Group 2 received letrozole (LET group); Group 3 received silymarin (SIL group) and Group 4 received no medication (CONT group). Experimental groups were treated for 3 weeks and then were sacrificed for volume and histopathological evaluation of implants and biochemical assessment. Serum and peritoneal levels of vascular endothelial growth factor (VEGF), total antioxidant activity (TAC) and tumor necrosis (TNF)-α were measured. RESULTS: Mean volume of the implants decreased significantly in silymarin (p < 0.001), letrozole (p < 0.001) and cabergoline (p < 0.001) groups compared to the control. Histopathologic score was significantly lower in silymarin (p: 0.039), letrozole (p: 0.017) and cabergoline (p < 0.001) groups compared to the control. Those receiving silymarin had significantly higher serum TAC compared to control after 21 days of therapy (p < 0.001). CONCLUSION: Silymarin, Letrozole, and Cabergoline administration resulted in decreased size and histopathologic grade of the induced endometrial lesions in a rat model. Silymarin appears to be a virtual novel therapeutic agent for treatment of endometriosis.

Molecular Detection and Genotypic Characterization of Toxoplasma gondii in Paraffin-Embedded Fetoplacental Tissues of Women with Recurrent Spontaneous Abortion.

BACKGROUND: Congenital toxoplasmosis is an important cause of spontaneous abortion worldwide. However, there is limited information on detection and genotypic characterization of Toxoplasma gondii (T. gondii) in women with recurrent spontaneous abortion (RSA). The aim of this study is the molecular detection and genotypic characterization of T. gondii in formalin-fixed, paraffin-embedded fetoplacental tissues (FFPTs) of women with RSA that have referred to the Avicenna Research Institute in Tehran, Iran. MATERIALS AND METHODS: This experimental research was undertaken on 210 FFPTs of women with RSA. The information of the patients was collected from the archives of Avicenna Research Institute in Tehran, Iran. After DNA extraction, the presence of T. gondii was examined by nested polymerase chain reaction targeting the GRA6 gene. Genotyping was performed on positive samples using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) that targeted the GRA6 and SAG3 genes. Sequencing was conducted on two GRA6 positive samples. RESULTS: T. gondii DNA was detected in 3.8% (8/210) of the samples. Genotyping showed that all positive samples belonged to type III of the T. gondii genotype. Sequencing two genomic DNAs of the GRA6 gene revealed 99% similarity with each other and 99-100% similarity with T. gondii sequences deposited in GenBank. There were six patients with histories of more than three abortions; one patient had a healthy girl and another patient had two previous abortions. Abortions occurred in the first trimester of pregnancy in seven patients and in the second trimester of pregnancy in one patient. CONCLUSION: The results of this study have indicated that genotype III is the predominant type of T. gondii in women with RSA in Tehran, Iran. Also, our findings suggest that toxoplasmosis may play a role in the pathogenesis of RSA. However, further studies are needed to elucidate a clear relationship between T. gondii infection and RSA.

Correlation between Gleason Scores in Needle Biopsy and Corresponding Radical Prostatectomy Specimens: A Twelve-Year Review.

BACKGROUND: Presence of discordance between the Gleason score on needle biopsy and the score of radical prostatectomy specimen is common and universal. In this study, we determined the accuracy of Gleason grading of biopsies in predicting histological grading of radical prostatectomy specimens and the degree of overgrading and undergrading of prostatic adenocarcinoma in our center, which is one of the referral centers in Tehran. METHODS: In this retrospective study, we analyzed the results of prostate needle biopsies and subsequent prostatectomies diagnosed at the Pathobiology Laboratory Center, Tehran, Iran in 45 patients between 2002 and 2013. Preoperative clinical data and the information from biopsy and prostatectomy specimens were collected. The accuracy, sensitivity, specificity, and positive and negative predictive values of different grades and groups were assessed. Pearson and Spearman correlation coefficient were used to determine the relation of different variables. RESULTS: The biopsy Gleason score was identical to the scores in prostatectomy specimens in 68.2% cases, while 31.8% were discrepant by 1 or 2 Gleason score. We had 9.1% downgrading and 22.7% cases upgraded after prostatectomy. The sensitivity and positive predictive value was 86% and 79% for low grade, 67% and 75% for moderate grade, and 80% and 80% for high-grade tumors, respectively. CONCLUSION: Overall, the reliability of Gleason grading of needle biopsies in predicting final pathology was satisfavory. Moderate grade group was the most difficult to diagnose in needle biopsy.

Molecular detection of Toxoplasma gondii in house sparrow (Passer domesticus) by LAMP and PCR methods in Tehran, Iran.

Toxoplasma gondii is one of the most common zoonotic parasitic diseases in human and warm-blooded animals worldwide. Birds are one of important intermediate hosts of T. gondii. The aim of this study is molecular detection of T. gondii in the house sparrow by LAMP and PCR methods in Tehran, Iran. A total 200 sparrows were captured in different regions of Tehran. DNA was extracted from tissue samples of each sparrow. LAMP and conventional PCR assays were carried out with a set of primers to detect the 529 bp fragment of T. gondii. LAMP and PCR were detected T. gondii from 17 (8.5 %) and 15 (7.5 %) of 200 sparrows respectively. These results indicated that sensitivity of LAMP was higher than conventional PCR. In our knowledge, this study is the first report of detection of T. gondii by LAMP method in bird hosts. Also, these findings provided an insight into epidemiological pattern of T. gondii infection in sparrow in Iran.

Breast cancer cells imaging by targeting methionine transporters with gadolinium-based nanoprobe.

PURPOSE: Early cancer diagnosis using MRI imaging is of high global interest as a non-invasive and powerful modality. In this study, methionine was conjugated on gadolinium-based mesoporous silica nanospheres to evaluate intra-cellular uptake and its accumulation in human breast cancer cells. PROCEDURES: The contrast agent was synthesized and characterized using different techniques including N2 physisorption, thermal gravimetric analysis, dynamic light scattering, and inductively coupled plasma atomic emission spectroscopy (ICP-AES). The intra-cellular uptake of Gd(3+) was measured by ICP-AES, fluorescent microscopy, and flow cytometry. Finally, cellular and tumor MR imaging were performed to determine in vitro and in vivo relaxometry. RESULTS: According to the results, the contrast agents accumulated in tumor cells both in vitro and in vivo. There was no significant cellular toxicity on either normal or cancer cells along with strong intense signal on T 1 compared to the unlabeled cells. CONCLUSIONS: The results showed that the novel contrast agent could become a useful tool in early detection of cancer.

Prevalence of intratubular germ cell neoplasia in cryptorchid testes of infertile men.

BACKGROUND: Cryptorchidism is a common malformation in neonates; surgery or medical treatments are applied during childhood. Untreated cryptorchid testes are in the risk of intratubular germ cell neoplasia (IGCN) and consequently invasive testicular tumors which could be shown by immunohistochemistry staining for placental like acid phosphatase (PLAP) marker. OBJECTIVE: We designed this study to know the prevalence of IGCN in untreated cryptorchid testes of infertile men, in our infertility center as a refferal center. MATERIALS AND METHODS: In this cross-sectional study we assessed H&E slides of testicular samples of 13 adult infertile patients with impalpable intra-abdominal testes seeking infertility treatment; then we stained them by PLAP marker. RESULTS: Three (23.08%) samples were positive for PLAP marker means presence of IGCN in testis. One of them showed seminoma besides IGCN. CONCLUSION: According to the results of this study and the fact that there are adult untreated cryptorchid patients in our country yet, it is suggested to pay more attention in clinical examination, assessment and follow up of such patients for malignancy screening.

Discrepancy in the frequency of Y chromosome microdeletions among Iranian infertile men with azoospermia and severe oligozoospermia.

AIM: There are discrepancies in the reported frequency of Y chromosome microdeletions among Iranian infertile men. The objective of this study was to determine the frequency of Y chromosome microdeletions in an Iranian population with azoospermia and severe oligozoospermia. METHODS: Totally, 94 azoospermic and 21 severe oligozoospermic patients were screened for the presence of Y chromosome microdeletions. One hundred and five fertile men were included as a control group, as well. The screening of Yq microdeletions was performed by two multiplex PCRs using six STS markers. RESULTS: No microdeletions were detected in the men with severe oligozoospermia. In the azoospermic group 2/94 (2.13%) patients showed Y chromosome microdeletions. Among them, one patient had complete deletion of the AZFc region and the other showed complete deletion of both the AZFb and AZFc regions (AZFbc). No microdeletion was identified in the AZFa region. CONCLUSION: The estimated frequency of Y chromosome microdeletions in the present survey was lower than that of previous reports in Iranian populations.

Seminal molecular markers as a non-invasive diagnostic tool for the evaluation of spermatogenesis in non-obstructive azoospermia.

Non-obstructive azoospermia (NOA) is currently evaluated by the use of conventional histopathological methods. In some cases, focal spermatogenesis is present in the testes of patients with NOA which may be almost undetectable by routine histopathological examinations. Application of molecular markers in semen to predict the spermatogenesis status in the testis will emphasize the probability of finding sperm in NOA testis through further search using TESE or mTESE. Detection of germ cell-specific transcripts in semen is a signal of germ cells present in the testis. In this study, we used molecular methods to evaluate spermatogenesis status in azoospermic men. Semen samples were collected from 203 men with azoospermia. Total RNA was extracted from the semen precipitates. First-strand complementary deoxyribonucleic acid (cDNA) was synthesized by reverse transcriptase then, (RT)-PCRs were carried out using primers for testis stage-specific genes (DAZ, AKAP4, PRM1, and PRM2). Testicular tissue biopsies were used for evaluating spermatogenesis status in testis. Histopathological examination and LH, FSH, and testosterone level measurements (chemiluminescence assay) were performed. The presence of DAZ and PRM2 transcripts in semen significantly indicated the presence of spermatogonia and spermatids in the testicular tissues. Absence of all four markers in semen confirmed the histopathological results corresponding to sertoli cell only syndrome (SCO). Although TESE should not be excluded solely on this criteria, using PRM1, PRM2, AKAP4, and DAZ transcripts in semen would provide a non-invasive molecular diagnostic tool to better counsel patients before undergoing TESE.

Production of Monoclonal Antibody against Human Nestin.

We have employed a peptide-based antibody generation protocol for producing antibody against human nestin. Using a 12-mer synthetic peptide from repetitive region of human nestin protein devoid of any N- or O-glyco-sylation sequences, we generated a mouse monoclonal antibody capable of recognizing human, mouse, bovine, and rat nestin. A wide variety of nestin proteins ranging from 140-250 kDa was detected by this antibody. This antibody is highly specific and functional in applications such as ELISA, flow cytometry, immunocytochemistry, and Western blot assays.

Ectopic Expression of Sortilin 1 (NTR-3) in Patients with Ovarian Carcinoma.

Gene expression profiling of ovarian carcinoma tissues has shown an increase of four-fold expression of SORT1 gene. Sortilin 1 (NTR-3) is a 95-100 kDa protein normally expressed in heart, brain, placenta, skeletal muscle, spinal cord, thyroid, and testis. However, its expression has never been reported in normal ovary. Here, we report expression of sortilin 1 in ovarian carcinoma tissues both at gene and protein levels. Sortilin 1 was expressed in all ovarian carcinoma patients (n=15) as well as ovarian carcinoma cell lines (n=5) regardless of their phenotypic characteristics. Non-malignant ovaries (n=6) did not express sortilin 1. The molecular basis for this ectopic expression is not yet clear. Our results showed a major cell surface expression of sortilin 1 rather than ER-Golgi compartment where it is mainly expressed. This finding may introduce sortilin 1 as a novel tumor marker for diagnosis of ovarian carcinoma and may signify its therapeutic value in targeted therapy.