Autophagy, a critical element in the aging male reproductive disorders and prostate cancer: a therapeutic point of view.

Autophagy is a highly conserved, lysosome-dependent biological mechanism involved in the degradation and recycling of cellular components. There is growing evidence that autophagy is related to male reproductive biology, particularly spermatogenic and endocrinologic processes closely associated with male sexual and reproductive health. In recent decades, problems such as decreasing sperm count, erectile dysfunction, and infertility have worsened. In addition, reproductive health is closely related to overall health and comorbidity in aging men. In this review, we will outline the role of autophagy as a new player in aging male reproductive dysfunction and prostate cancer. We first provide an overview of the mechanisms of autophagy and its role in regulating male reproductive cells. We then focus on the link between autophagy and aging-related diseases. This is followed by a discussion of therapeutic strategies targeting autophagy before we end with limitations of current studies and suggestions for future developments in the field.

Comparison of Rabies Cases Received by the Shomal Pasteur Institute in Northern Iran: A 2-Year Study.

The rabies virus, which belongs to the genus Lyssavirus, the family Rhabdoviridae, is the causative agent of rabies, a contagious, deadly, and progressive neurological infection. This illness is commonly distributed worldwide and affects all warm-blooded animals. Regarding the zoonotic aspects of rabies, the prevalence of rabies was investigated in this study. Over 2 years, 188 samples were examined via the direct fluorescent antibody test (DFAT) and mouse inoculation test (MIT) techniques by using brain tissue samples. Our findings showed that 73.94% of samples were rabies positive. The highest number of samples belonged to cows and dogs, respectively. The positivity rate in cows was 71.88%, followed by dogs with a 57.78% infection rate. These findings suggested that despite the heavy monitoring protocols in Iran, rabies is still a prevalent disease, and it is advised that vaccinations and screening programs should be carried out more frequently with heavier observation.

Embryonic stem cell differentiation to primordial germ cell like cells by Nigella sativa, Brassica Oleracea and Oenothera biennis extracts.

OBJECTIVES: This study aimed to investigate the induction effects of methanolic extracts of Nigella sativa (NiS), Brassica Oleracea (BrO), and Oenothera biennia (Obi) on transgenic embryonic stem cells (ESCs) and to evaluate the ability of germ cells (GCs) production using these pluripotent cells. METHODS: ESCs were amplified using a feeder layer. Embryoid bodies enzymatically dissociated to single cells and induced the extracts in gelatinized plates. Then RNA extraction and cDNA synthesis were performed. In the presence of appropriate primers, the desired genes were quantitatively evaluated by quantitative polymerase chain reaction (qPCR). RESULTS: The copies of all genes in the control group showed a decreasing trend during the first to third weeks. Compared to the control group, the expression level of sex determining region Y-box 2 gene (Sox2) showed the highest level. All four evaluated genes increased in all Obi groups compared to the control group. There is also a slight increase in the Nanog homeobox gene (Nanog). Obi extract in different concentrations has increased the expression of the Sox2 gene. Increased expression of this gene along with octamer-binding transcription factor 4 gene (Oct4) and Nanog indicates a condition close to germ cell-like cells (GCLCs). CONCLUSIONS: According to the results of this study, NiS can increase expression of the Oct4, Sox2, Nanog, and stimulated by retinoic acid gene 8 (STRA8) genes and so increase the hope of GCs production. Storage of cells for 21 days in the presence of the extract compared to 14 days has a negative effect on cell growth and differentiation. The effects of meiosis onset and GCs production can be expected in the presence of some herbal extracts. Optimal utilization of these extracts requires further study in the field of different extracts and fractions of each extract to more effectively and purposefully direct the differentiation of stem cells.

Long-time effects of prenatal morphine, tramadol, methadone, and buprenorphine exposure on seizure and anxiety in immature rats.

Purpose: This study aimed to investigate seizures and anxiety-like behaviors in immature rats prenatally exposed to opiate drugs.Materials and methods: Pregnant rats were randomly divided into five groups: saline, morphine, tramadol, methadone, and buprenorphine. Administrations were performed intraperitoneally once a day for the last seven days of pregnancy. Neonatal rats were subdivided into ten groups, split according to sex. Anxiety-like behavior was tested on postnatal day (PD) 19. On PD 20, seizure was induced by PTZ injection.Results: Morphine in male rats had an increased time to onset (p < 0.005), whereas there was a decreased number of tonic-clonic seizures in females (p < 0.05). Tramadol had an increased duration of tonic-clonic seizures compared to morphine and methadone in males (p < 0.005). Moreover, tramadol decreased open arm time and locomotor activity in males more than in females (p < 0.05). Methadone decreased open arm time in males (p < 0.05). Furthermore, buprenorphine and tramadol decreased open arm entrance in male rats (p < 0.05).Conclusions: It was demonstrated that prenatal tramadol significantly increases both the duration of seizures and anxiety-like behaviors in immature male rats, whereas morphine decreases both of them. The effects of tramadol on seizure and anxiety-like behavior may be due to the comorbid occurrence of the symptoms of these two disorders.

Comparison of Germ Cell Gene Expressions in Spontaneous Monolayer versus Embryoid Body Differentiation of Mouse Embryonic Stem Cells toward Germ Cells.

BACKGROUND: Genetic and morphologic similarities between mouse embryonic stem cells (ESCs) and primordial germ cells (PGCs) make it difficult to distinguish differentiation of these two cell types in vitro. Using specific GC markers expressed in low level or even not expressed in ESCs- can help recognize differentiated cells in vitro. We attempted to differentiate the mouse ESCs into GC-like cells spontaneously in monolayer and EB culture method. MATERIALS AND METHODS: In this experimental study, we attempted to differentiate ESCs, Oct4-GFP OG2, into GC-like cells (GCLCs) spontaneously in two different ways, including: i. Spontaneous differentiation of ESCs in monolayer culture as (SP) and ii. Spontaneous differentiation of ESCs using embryoid body (EB) culture method as (EB+SP). During culture, expression level of four GC specific genes (Fkbp6, Mov10l1, Riken and Tex13) and Mvh, Scp3, Stra8, Oct4 were evaluated. RESULTS: In both groups, Mov10l1 was down-regulated (P=0.3), while Tex13 and Riken were up-regulated (P=0.3 and P=0.04, respectively). Fkbp6 and Stra8 were decreased in EB+SP and they were increased in SP group, while no significant difference was determined between them (P=0.1, P=0.07). Additionally, in SP group, gene expression of Mvh and Scp3 were up-regulated and they had significant differences compared to EB+SP group (P=0.00 and P=0.01, respectively). Oct4 was down-regulated in the both groups. Flow-cytometry analysis showed that mean number of Mvh-positive cells in the SP group was significantly greater compared to ESCs, EB+SP and EB7 groups (P=0.00, P=0.01, and P=0.3, respectively). CONCLUSION: These findings showed that ESCs were differentiated into GCLCs in both group. But spontaneous differentiation of ESCs into GCLCs in SP group (monolayer culture) compared to EB+SP (EB culture methods) has more ability to express GCs markers.

Evaluation of specific germ cell genes expression in mouse embryonic stem cell-derived germ cell like cells treated with bone morphogenetic protein 4 in vitro.

BACKGROUND: Bone morphogenetic protein 4 (BMP4) is a significant signaling molecule that involves in initiating of differentiation and performs multifunctional effects on embryonic stem cells (ESCs) and embryos. OBJECTIVE: The goal of the present study was to evaluate an in vitro differentiation model of mouse embryonic stem cells into germ cells, using BMP4. MATERIALS AND METHODS: in this experimental study, we used Oct4-GFP mouse ESCs to form embryoid body (EB) aggregations for two days. Then, single cells from EB were cultured for four days with BMP4. Using MTT assay and gene expression levels for evaluation of Mvh and Riken by real-time RT-PCR of six concentrations, 12.5 ng/ ml BMP4 was determined as an optimized dose. Then, the expression level of Fkbp6, Mov10l1, 4930432K21Rik, Tex13, Mvh, Scp3, Stra8, Oct4 were evaluated. Flow cytometry and immunostaning were used to confirm the findings of the real-time RT-PCR. RESULTS: In the +BMP4 group, the genes encoding Riken (p≤0.001) and Mvh (p≤0.001) were found to be increased with significant differences compared with the control group. Mov10l1 (p=0.22), Tex13 (p=0.10), Fkbp6 (p=0.90), Scp3 (p=0.61) and Stra8 (p=0.08) were up-regulated without significance differences compared with control group. Flow cytometry analysis showed that the mean number of Mvh-positive cells in the +BMP4 group was greater when compared with ESCs, -BMP4 and EB groups (p=0.03, p≤0.001, p=0.02, respectively). CONCLUSION: Down-regulation of Oct4, expression of germ cells genes and meiosis markers expression raise this hypothesis that ESCs were differentiated by BMP4, and may be introduced into the first meiosis as germ cell-like cells.

Evaluation of Novel Mouse-Specific Germ Cell Gene Expression in Embryonic Stem Cell-Derived Germ Cell-Like Cells In Vitro with Retinoic Acid Treatment.

We designed a study to induce differentiation of Oct4-GFP (expression of Green Fluorescent Protein of oct4) embryonic stem cells (ESCs) by embryoid body (EB) culture system into germ cells (GCs) using retinoic acid (RA) and evaluated the expression level of (Fkbp6, Mov10l1, 4930432K21Rik, and Tex13) in differentiated cells. The expression levels of four GC-related genes, Oct4, Mvh, Scp3, and Stra8, was determined by quantitative real-time polymerase chain reaction (q-RT-PCR). Immunostaining and flow cytometry were used as additional tests to confirm q-RT-PCR findings. A significant increase occurred in the expression of meiotic markers and specific genes, Fkbp6 (p = 0.00), Mov10l1 (p = 0.01), and Tex13 (p = 0.00) in ESCs treated with RA (+RA) compared with the controls (-RA). Oct4 expression was decreased in all studied groups. The expression levels of 4930432K21Rik, Mvh, Stra8, and Scp3 in the +RA group was higher than that of the -RA group. Flow cytometry analysis showed that mean number of Mvh-positive cells in the +RA group was greater as compared with ESCs, -RA and EB7 groups (p = 0.00). Downregulation of Oct4 as a pluripotency factor as well as the expression of meiosis markers, this hypothesis is raised that ESCs are differentiated by RA, and have been introduced into the zygote/pachytene of first meiosis as GC-like cells.

In vitro and in vivo evaluation of therapy targeting epithelial-cell adhesion-molecule aptamers for non-small cell lung cancer.

Targeted, disease-specific delivery of therapeutic nanoparticles shows wonderful promise for transmitting highly cytotoxic anti-cancer agents. Using the reaction of non-small cell lung cancer (SK-MES-1 and A549 cell lines) as representative of other cancer types', the present study examines the effects of EpCAM-fluoropyrimidine RNA aptamer-decorated, DOX-loaded, PLGA-b-PEG nanopolymersomes that bond specifically to the extracellular domain of epithelial-cell adhesion molecules. Results demonstrate that EpCAM aptamer-conjugated DOX-NPs (Apt-DOX-NP) significantly enhance cellular nanoparticle uptake in SK-MES-1 and A549 cell lines and increase the cytotoxicity of the DOX payload as compared with non-targeted DOX-NP (P<0.05). Additionally, Apt-DOX-NP exhibits greater tumor inhibition in nude mice bearing SK-MES-1 non-small cell lung-cancer xenografts and reduces toxicity, as determined by loss of body weight, cardiac histopathology and animal survival rate in vivo. After a single intravenous injection of Apt-DOX-NP and DOX-NPs, tumor volume decreased 60.9% and 31.4%, respectively, in SK-MES-1-xenograft nude mice compared with members of a saline-injected control group. This study proves the potential utility of Apt-DOX-NP for therapeutic application in non-small cell lung cancer. In the future, EpCAM-targeted therapies might play a key role in treating non-small cell lung cancer, the most common type of lung cancer.

α:Non-α and Gγ:Aγ globin chain ratios in thalassemia intermedia patients treated with hydroxyurea.

OBJECTIVES: To elucidate the possible ways by which hydroxyurea molecules affect globin chain (α or ß-like) synthesis. METHODS: A total of 23 thalassemia intermedia patients (13 male and 10 female) aged between 5 and 26 years were treated for five months with 15 mg/(kg·day) of hydroxyurea. Hemoglobins electrophoresis and globin chain electrophoresis was performed on each sample at different time points before and during the treatment. RESULTS: Fetal hemoglobin increased significantly in most patients and average episodes of transfusion decreased. Both Gγ and Aγ-globin chains increased significantly and α-globin:Nonα-globin chain as well as Gγ-globin:Aγ globin chains ratios decreased. CONCLUSIONS: Improvement in α:non-α ratio and consequent decrease of free α-globin chain might be the cause of beneficial effects of hydroxyurea therapy. Two patients who felt better didn't show significant increase in their fetal hemoglobin level, and this is in contradiction with the hypothesis claiming that the HbF level increase is the cause of such therapeutic effect. In spite of the unclear mechanism of action of this drug, hydroxyurea therapy had noticeable impacts on thalassemia intermedia and also sickle cell disease and even patients suffering from thalassemia major.