Effects of cigarette smoke condensate on reproduction in mice in vivo.

Smoking has dangerous and sometimes irreversible effects on various body tissues, including the reproductive system. We conducted this research to determine the in vivo effects of cigarette smoke condensate (CSC) on reproduction in mice. In this experimental in vivo study, 32 male and female NMRI mice were divided into four groups. The mice were injected with CSC (CSC-1R3F) for 28 days. The mice were mated 1 day after the last injection and observed daily for 1 week for the presence of a vaginal plug to track mating. We evaluated mating success rate, and sperm and oocyte quality, pregnancy outcome, childbearing status, and in vitro fertilization (IVF). The results showed a decrease in successful mating in female mice that received the CSC injections. CSC significantly influenced the number of offspring born to males. When the CSC was injected into male mice, there was a significant increase in the number of offspring compared with the group in which only the females received CSC injections. According to the results, there was a negative effect of CSC on morphological parameters in male and female mice. Also, successful IVF after exposure to CSC was significantly decreased in the female mice treated group. The results indicated that CSC significantly affected the number of offspring and fecundity success in females.

Osteogenic effect of electromagnetic fields on stem cells derived from rat bone marrow cultured in osteogenic medium versus conditioned medium in vitro.

OBJECTIVES: This study assessed possible osteogenic differentiation caused by electromagnetic fields (EMF) on rat bone-marrow-derived stem cells (rBMSCs) cultured in osteogenic medium (OM) or in human adipose-stem cell-conditioned medium (hADSC-CM). MATERIALS AND METHODS: The rBMSCs were divided into negative and positive control groups, cultured in α-MEM plus 10% FBS or OM respectively. CM and CM + EMF  groups, cultured cells in hADSCs-CM or exposed to EMF (50 Hz, 1 mT) for 30 min/day plus hADSCs-CM, respectively. Cells from the OM + EMF were simultaneously cultured in OM and exposed to EMF. Osteogenesis was investigated through alkaline phosphatase activity, alizarin red staining and real-time PCR. RESULTS: A meaningfully higher level of ALP activity was observed in the OM + EMF group compared to the other groups. There was a considerable increase in Runx2 expression in the CM + EMF group compared to the positive control and CM groups and a significant increase in Runx2 expression in the OM + EMF in comparison with all other groups after 21 days. Runx2 expression increased significantly in the CM, CM + EMF and positive control groups on day 21 compared to the same groups on day 14. From days 14-21, Ocn expression increased in the CM and CM + EMF groups, but both groups showed a significant decrease compared to the positive controls. CM and EMF had no effect on Ocn expression. On day 21, Ocn expression was significantly higher in the OM + EMF group than in the positive control group. CONCLUSION: The synergistic effect of EMF and OM increased the expression of Runx2 and Ocn in rBMSCs.

Feeding role of mouse embryonic fibroblast cells is influenced by genetic background, cell passage and day of isolation.

Mouse embryonic fibroblast (MEF) cells are commonly used as feeder cells to maintain the pluripotent state of stem cells. MEFs produce growth factors and provide adhesion molecules and extracellular matrix (ECM) compounds for cellular binding. In the present study, we compared the expression levels of Fgf2, Bmp4, ActivinA, Lif and Tgfb1 genes at the mRNA level and the level of Fgf2 protein secretion and Lif cytokine secretion at passages one, three and five of MEFs isolated from 13.5-day-old and 15.5-day-old embryos of NMRI and C57BL/6 mice using real-time PCR and enzyme-linked immunosorbent assay. We observed differences in the expression levels of the studied genes and secretion of the two growth factors in the three passages of MEFs isolated from 13.5-day-old and 15.5-day-old embryos, respectively. These differences were also observed between the NMRI and C57BL/6 strains. The results of this study suggested that researchers should use mice embryos that have different genetic backgrounds and ages, in addition to different MEF passages, when producing MEFs based on the application and type of their study.

Use of alginate hydrogel to improve long-term 3D culture of spermatogonial stem cells: stemness gene expression and structural features.

The quality and quantity of a spermatogonial stem-cell (SSC) culture can be measured in less time using a 3D culture in a scaffold. The present study investigated stemness gene expression and the morphological and structural characterization of SSCs encapsulated in alginate. SSCs were harvested from BALB/c neonatal mice testes through two-step mechanical and enzymatic digestion. The spermatogonial populations were separated using magnetic-activated cell sorting (MACS) using an anti-Thy1 antibody and c-Kit. The SSCs then were encapsulated in alginate hydrogel. After 2 months of SSC culturing, the alginate microbeads were extracted and stained to evaluate their histological properties. Real-time polymerase chain reaction (PCR) was performed to determine the stemness gene expression. Scanning electron microscopy (SEM) was performed to evaluate the SSC morphology, density and scaffold structure. The results showed that encapsulated SSCs had decreased expression of Oct4, Sox2 and Nanos2 genes, but the expression of Nanog, Bcl6b and Plzf genes was not significantly altered. Histological examination showed that SSCs with pale nuclei and numerous nucleolus formed colonies. SEM evaluation revealed that the alginate scaffold structure preserved the SSC morphology and density for more than 60 days. Cultivation of SSCs on alginate hydrogel can affect Oct4, Sox2 and Nanos2 expression.

Evaluation of co-cultured spermatogonial stem cells encapsulated in alginate hydrogel with Sertoli cells and their transplantation into azoospermic mice.

An in vitro spermatogonial stem cell (SSC) culture can serve as an effective technique to study spermatogenesis and treatment for male infertility. In this research, we compared the effect of a three-dimensional alginate hydrogel with Sertoli cells in a 3D culture and co-cultured Sertoli cells. After harvest of SSCs from neonatal mice testes, the SSCs were divided into two groups: SSCs on a 3D alginate hydrogel with Sertoli cells and a co-culture of SSCs with Sertoli cells for 1 month. The samples were evaluated by quantitative reverse transcription polymerase chain reaction (qRT-PCR) assays and bromodeoxyuridine (BrdU) tracing, haematoxylin and eosin (H&E) and periodic acid-Schiff (PAS) staining after transplantation into an azoospermic testis mouse. The 3D group showed rapid cell proliferation and numerous colonies compared with the co-culture group. Molecular assessment showed significantly increased integrin alpha-6, integrin beta-1, Nanog, Plzf, Thy-1, Oct4 and Bcl2 expression levels in the 3D group and decreased expression levels of P53, Fas, and Bax. BrdU tracing, and H&E and PAS staining results indicated that the hydrogel alginate improved spermatogenesis after transplantation in vivo. This finding suggested that cultivation of SSCs on alginate hydrogel with Sertoli cells in a 3D culture can lead to efficient proliferation and maintenance of SSC stemness and enhance the efficiency of SSC transplantation.

Effect of cigarette smoke condensate on mouse embryo development and expression of pluripotency and apoptotic genes in vitro.

The aim of the present study was to investigate the effect of cigarette smoke condensate (CSC) on in vitro development of mouse embryos. In total 3000 NMRI mice 2PN embryos were divided into six groups (n = 500). The test group was exposed to 20, 40, 80, 160 or 320 µg/ml of CSC. In the control group, CSC was not added to the culture medium during the development of 2PN embryos. The effects of 20 and 80 µg/ml of CSC on genes involved in pluripotency and apoptosis, and also, the aryl hydrocarbon receptor gene was assessed in the blastocysts. Our results showed that CSC had an adverse effect on the viability of mouse embryos at the concentrations of 80, 160 and 320 µg/ml compared with the control group (P < 0.05). In contrast, it had positive effects on the viability of mouse embryos at the concentrations of 20 and 40 µg/ml compared with the control group (P < 0.05). The 20 and 80 µg/ml concentrations of CSC increased the expression of pluripotency, apoptotic, and aryl hydrocarbon receptor genes in the blastocyst embryo stage compared with the control group (P < 0.05). It can be concluded that concentrations higher than 40 µg/ml of CSC have an adverse effect on mouse embryo development in the preimplantation stages. Also, 20 and 80 µg/ml concentrations of CSC have a significant effect on the expression of pluripotency, apoptotic, and the aryl hydrocarbon receptor genes in the blastocyst embryo stage compared with the control group.

Effect of cyanocobalamin on oocyte maturation, in vitro fertilization, and embryo development in mice.

The aim of this study was to investigate the effect of cyanocobalamin supplementation on in vitro maturation (IVM), in vitro fertilization (IVF), and subsequent embryonic development competence to the blastocyst stage, and in vitro development of mouse 2-cell embryos. Cumulus cells were prepared from mouse cumulus-oocyte complexes (COCs) and incubated for 24 h in an in vitro culture (IVC) medium that contained different concentrations of cyanocobalamin (100, 200, 300 or 500 pM). We collected 2-cell embryos from superovulated NMRI mice and cultured them in the same concentrations of cyanocobalamin (100, 200, 300 or 500 pM). After 42 h of IVM, we observed significantly increased oocyte maturation in the 200 pM cyanocobalamin-treated group compared with the control group (P < 0.0001). Mature oocytes cultured in 200 pM cyanocobalamin were fertilized and cultured in IVC medium with cyanocobalamin (100, 200, 300 or 500 pM) during early embryogenesis. The matured oocytes that were cultured in 200 pM cyanocobalamin had significantly higher 2-cell development rates compared with the control oocytes (P < 0.01). Embryos obtained from in vitro mature oocytes and in vivo fertilized oocytes that were cultured in 200 pM cyanocobalamin had significantly greater frequencies of development to the blastocyst stage and a significant reduction in 2-cell blocked and degenerated embryos compared with the control embryos (P < 0.0001). Embryos derived from oocytes fertilized in vivo with 200 pM cyanocobalamin had a higher percentage of blastocyst embryos compared with those derived from matured oocytes cultured in vitro (P < 0.0001). These finding demonstrated that the effects of cyanocobalamin on oocyte maturation, fertilization, and embryo development in mice depend on the concentration used in IVC medium.

Quercetin postconditioning attenuates gastrocnemius muscle ischemia/reperfusion injury in rats.

Quercetin, an antioxidant derived from plants, can play a beneficial role in the protection of various tissues against ischemia-reperfusion injuries (IRI). The purpose of the present research was to investigate the protective effects of quercetin on gastrocnemius muscle ischemia-reperfusion. A total of 80 adult male Wistar rats (weights: 250-300 g) were divided into ten groups (n = 8 per group). We used silk 6.0 surgical thread to create a knit to occlude the femoral artery and vein for 3 hr. The treated groups, which comprised half of each experimental group, received intraperitoneal injections of 150 mg/kg quercetin after the ischemia. Blood flow was subsequently reestablished in the reperfusion phase. The rats were kept in reperfusion for 3, 7, 14, or 28 days after which they were killed with high doses of anesthetic drugs, and the gastrocnemius muscles were removed and fixed. Tissue processing, hematoxylin and eosin and toluidine blue staining, and immunohistochemistry were used to assess tumor necrosis factor-α (TNF-α) and nuclear factor κB (NF-κB) levels. A comparison between treated and untreated ischemic sites showed that on the third day of reperfusion, the severity of edema and NF-κB level decreased significantly; on the 7th day of reperfusion, the severity of edema and the levels of TNF-α and NF-κB decreased significantly; and on the 14th day of reperfusion, all of the parameters showed significant decreases. On the 28th day of reperfusion, there were significantly decreased levels of TNF-α and NF-κB, and decreased mast cell infiltration when compared with the untreated groups. According to the results, administration of quercetin after ischemia could significantly prevent gastrocnemius muscle IRI.

A comparison of the effects of fetal bovine serum and newborn calf serum on cell growth and maintenance of cryopreserved mouse spermatogonial stem cells.

Serum is a common supplement that is widely used to protect various cells and tissues from cryopreservation because it provides the necessary active components for cell growth and maintenance. In this study, we compared the effects of newborn calf serum (NCS) and fetal bovine serum (FBS) on the cryopreservation of mouse spermatogonial stem cells (SSCs). The isolated SSCs were cryopreserved in two groups: freezing medium that contained 10% DMSO (dimethyl sulfoxide) and 10% FBS in DMEM (Dulbecco's Modified Eagle's Medium) (group 1) and freezing medium that contained 10% DMSO and 10% NCS in DMEM (group 2). Real-time PCR was performed for stemness gene expression. The SSCs' viability was performed by trypan blue. We observed that the SSCs had increased viability in the NCS-freeze/thaw group (87.82%) compared to the FBS-freeze/thaw group (79.83%), but this increase was not statistically significant (P < 0.105). Promyelocytic leukemia zinc finger (Plzf) and Lin28 gene expression levels in the NCS-frozen/thawed SSCs were not significantly different compared to the FBS-frozen/thawed SSCs; however, Nanog gene expression increased considerably, and Dazl gene expression decreased significantly. The results in this study demonstrated that the presence of NCS in a solution of cryopreserved SSCs increased their viability after freeze/thawing and might promote the proliferation of cultivated SSCs in vitro by increasing the relative expression of Nanog.

Protective effects of royal jelly on testicular torsion induced ischaemia reperfusion injury in rats.

This study was performed to investigate the protective effects of royal jelly (RJ) on a testicular torsion-induced ischaemia/reperfusion (I/R) injury in adult rats. A total of 40 male Wistar rats were divided into four groups, including 10 rats in each group: Group 1 (sham), Group 2 (Control), group 3 (I/R rats treated with 100 mg/kg RJ for 50 days after torsion) and group 4( I/R rats treated with 20 mg/kg vitamin C for 50 days after torsion). Testicular torsion was created by rotating the right testes 720° a clockwise direction for 90 min. The levels of testosterone were measured by ELISA. Pathological evaluation, mean maturity and quality of the seminiferous tubules were used. Results showed that the testicular histopathology standards and testosterone levels changes were statistically significant in groups 3 and 4. The results obtained in this study may suggest that RJ like vitamin C had protective effects on a testicular ischaemia/reperfusion-induced injury in rats.

Effect of embryo cryopreservation on derivation efficiency, pluripotency, and differentiation capacity of mouse embryonic stem cells.

Mouse embryonic stem cells (mESCs) are pluripotent cells that have the capability for self-renewal. One of the most important factors that affect the efficiency of their isolation is the condition of the mouse embryos. The main objective of this study is to isolate mESCs from C57BL/6 frozen/thawed eight-cell mouse embryos using serum-free culture. We generated mESCs from blastocysts that developed from frozen/thawed embryos of C57BL/6 mice by the 3i + LIF medium. Assessments of the isolated mESC lines (MUKF-1, MUKF-2, and MUKF-3) included simple karyotype analysis; polymerase chain reaction of the testis-determining gene (Sry); determination of alkaline phosphatase (ALP) activity; expressions of pluripotent transcription factors Oct4, Rex1, Sox2, and Nanog by reverse transcription polymerase chain reaction; and immunocytochemistry assessment of OCT4 and SSEA-I expressions at the protein level. We evaluated the ability of these mESC lines to differentiate into three germ layers by embryoid body (EB) formation. The cell doubling time (DT) of isolated mESCs was determined. The 2-C57 cell line was served as control. Germline competence of the male mESC line (MUKF-3) was tested through chimeric mouse production. Three independent mESC lines (MUKF-1, MUKF-2, and MUKF-3) were established from five cryopreserved embryos. The MUKF-1 and MUKF-2 lines were female, whereas MUKF-3 was a male mESC line. Karyotype analysis showed that MUKF-3 had a diploid karyotype, whereas MUKF-1 and MUKF-2 had abnormal karyotypes. All three lines had ALP activity and expressed Oct4, Rex1, and Nanog. Immunocytochemistry assessment for OCT4 and SSEA-I was positive for all three lines. The DT differed in the three mESC lines. MUKF-1 and MUKF-3 could form EB and express developmental genes after spontaneous differentiation. These data demonstrated that probably cryopreservation affected the efficiency of derivation, karyotype, DT, expression of pluripotency, developmental genes, and differentiation capacity of the independent mESC lines.

Interaction and molecular dynamics simulation study of Osimertinib (AstraZeneca 9291) anticancer drug with the EGFR kinase domain in native protein and mutated L844V and C797S.

BACKGROUND: Targeted therapy is a novel, promising approach to anticancer treatment that endeavors to overcome drug resistance to traditional chemotherapies. Patients with the L858R mutation in epidermal growth factor receptor (EGFR) respond to the first generation tyrosine kinase inhibitors (TKIs); however, after one year of treatment, they may become resistant. The T790M mutation is the most probable cause for drug resistance. Third generation drugs, including Osimertinib (AZD9291), are more effective against T790M and other sensitive mutations. Osimertinib is effective against the L844V mutation, has conditional effectiveness for the L718Q mutation, and is ineffective for the Cys797Ser (C797S) mutation. Cells that have both the T790M and C797 mutations are more resistant to third generation drugs. Although research has shown that Osimertinib is an effective treatment for EGFR L844V cells, this has not been shown for cells that have the C797S mutation. This molecular mechanism has not been well-studied. METHODS: In the present study, we used the GROMACS software for molecular dynamics simulation to identify interactions between Osimertinib and the kinase part of EGFR in L844V and C797S mutants. RESULTS: We evaluated native EGFR protein and the L844V and C797S mutations' docking and binding energy, kI, intermolecular, internal, and torsional energy parameters. Osimertinib was effective for the EGFR L844V mutation, but not for EGFR C797S. All simulations were validated by root-mean-square deviation (RMSD), root-mean square fluctuation (RMSF), and radius of gyration (ROG). CONCLUSION: According to our computational simulation, the results supported the experimental models and, therefore, could confirm and predict the molecular mechanism of drug efficacy.

Increasing maternal age of blastocyst affects on efficient derivation and behavior of mouse embryonic stem cells.

Mouse embryonic stem cells (mESCs) have the capability to undergo unlimited cell division and differentiate into derivatives of all three embryonic germ layers. These fundamental features enable mESCs to potentially be appropriate, efficient models for biological and medical research. Therefore, it is essential to produce high-performance mESCs. In the current study, we have produced mESCs from blastocysts that developed from fertilized oocytes of 2 (2-C57)-, 4 (4-C57)-, and 6 (6-C57)-month-old C57BL/6 mice. A comparison of isolated stem cells was done from the viewpoint of the efficiency of mESC derivation, self-renewal, and their differentiation capacity. All generated mESCs showed a similar expression of the molecular markers protein of pluripotency and AP activity. In the 3i medium, there was a significant decrease in undifferentiated marker genes expression in the 2-C57 cells compared with the other two groups ( P < 0.05) but developmental genes significantly increased in the 4-C57 and 6-C57 cells compared with the 2-C57 cells ( P < 0.05). The differentiation capacity into three germ layers through the embryoid body formation and percentage of cell lines with normal numbers of chromosomes reduced with increased maternal age. The highest DT and highest percentage of cells in the S phase belonged to 2-C57 cells. These data demonstrated that blastocysts which developed from fertilized oocytes of 2-, 4-, and 6-month-old C57BL/6 mice can generate pluripotent stem cells, and suggested that both the efficiency of mESC isolation and the behavior of these isolated mESCs including pluripotency, self-renewal, cell cycle, and DT changed with increasing maternal age.

Effects of cigarette smoke condensate on proliferation and pluripotency gene expression in mouse embryonic stem cells.

BACKGROUND: Embryonic stem cells (ESCs) are derived from the inner cell mass (ICM) of blastocysts. They can be used as valuable experimental models to test the effects of drugs, chemicals, and environmental contaminants such as cigarette smoke condensate (CSC) on preimplantation embryo development. The aim of this study was to evaluate the effect of CSC on ESCs derived from mice with different genetic backgrounds and maternal ages. METHODS: The study groups consisted of mouse ESCs (mESCs) obtained from three sources: blastocysts developed from fertilized oocytes of two-month-old (2-C57) and six-month-old (6-C57) C57BL/6 inbred mice and those developed from fertilized oocytes of two-month-old (2-NMRI) NMRI outbred mice. The groups of mESCs were exposed to 0.04, 4, and 40 µg/mL CSC. After exposure, we measured cell viability by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay and real-time polymerase chain reaction for changes in expressions of Oct4, Sox2, Nanog, Ahr, Bax, Bcl2, TFAM, and POLG. The cell doubling time (DT) of these populations was also determined. RESULTS: We observed that CSC changed proliferation and DT in the 2-C57 and 6-C57 cells. There was no change in 2-NMRI cells. Exposure to CSC caused changes in the gene expressions and induced apoptosis in all three cell lines. CONCLUSION: Based on the results of the study, it can be concluded that CSC has an effect on the viability, DT and gene expression patterns in mouse ESCs and its effects vary based on the genetic background and maternal age of isolated mouse ESCs.

Generation of Fam83h knockout mice by CRISPR/Cas9-mediated gene engineering.

Family with sequence similarity 83 member H (FAM83H) protein-coding geneplay an essential role in the structural organization, calcification of developing enamel, and keratin cytoskeleton disassembly by recruiting Casein kinase 1 alpha (CSNK1A1) to keratin filaments. In this study, we have applied CRISPR Cas9 nickase (D10A) to knockout (KO) the Fam83h gene in NMRI outbred mice. We generated homozygous Fam83h KO mice ( Fam83h Ko/Ko ) through a premature termination codon, which was validated by Sanger sequencing in F0 generation. Next, we also bred the FAM83H KO for two generations. Reverse-transcription polymerase chain reaction and Western blot analysis approved the Fam83h KO mice. The Fam83h KO mice had evidence of normal morphology at the cervical loops, secretory and maturation stages, and mandibular molars. In comparison with the normal wild-type mice ( Fam83h W/W ), the F2 homozygous KO ( Fam83h Ko/Ko ) had sparse, scruffy coats with small body size and decreased general activity. Also, they had the natural reproductive ability and natural lifespan. In addition, delay in opening the eyes and dry eyes among infant mice were seen. The F1 heterozygous mice looked comparable to the normal wild-type mice ( Fam83h W/W ), which showed autosomal recessive inheritance of these phenotypes. The KO of FAM83H had controversial effects on the development of teeth and the formation of enamel. The phenotype defect in dental development and the enamel formation were seen in three mice among four generations. It can be concluded that null FAM83H in outbred mice not only showed the reported phenotypes in null inbred mouse but also showed normal lifespan and reproductive ability; dental deficiency in three homozygous mice; and the symptoms that were similar to the symptoms of dry eye syndrome and curly coat dog syndrome in all four evaluated KO generations.

Intracranial Hemorrhage: A Devastating Outcome of Congenital Bleeding Disorders-Prevalence, Diagnosis, and Management, with a Special Focus on Congenital Factor XIII Deficiency.

Intracranial hemorrhage (ICH) is a medical emergency. In congenital bleeding disorders, ICH is a devastating presentation accompanied with a high rate of morbidity and mortality. The prevalence of ICH is highly variable among congenital bleeding disorders, with the highest incidence observed in factor (F) XIII deficiency (FXIIID) (∼30%). This life-threatening presentation is less common in afibrinogenemia, FVIII, FIX, FVII, and FX deficiencies, and is rare in severe FV and FII deficiencies, type 3 von Willebrand disease and inherited platelet function disorders (IPFDs). In FXIIID, this diathesis most often occurs after trauma in children, whereas spontaneous ICH is more frequent in adults. About 15% of patients with FXIIID and ICH die; the bleeding causes 80% of deaths in this coagulopathy. Although in FXIIID, the bleed most commonly is intraparenchymal (> 90%), epidural, subdural, and subarachnoid hemorrhages also have been reported, albeit rarely. As this life-threatening bleeding causes neurological complications, early diagnosis can prevent further expansion of the hematoma and secondary damage. Neuroimaging plays a crucial role in the diagnosis of ICH, but signs and symptoms in patients with severe FXIIID should trigger replacement therapy even before establishment of the diagnosis. Although a high dose of FXIII concentrate can reduce the rate of morbidity and mortality of ICH in FXIIID, it may occasionally trigger inhibitor development, thus complicating ICH management and future prophylaxis. Nevertheless, replacement therapy is the mainstay of treatment for ICH in FXIIID. Neurosurgery is performed in patients with FXIIID and epidural hematoma and a hemorrhage diameter exceeding 2 cm or a volume of ICH is more than 30 cm3. Contact sports are not recommended in people with FXIIID as they can elicit ICH. However, a considerable number of safe sports and activities have been suggested to have more benefits than dangers for patients with congenital bleeding disorders, and are hence suitable for these patients.

Selenium mitigates methotrexate-induced testicular injury: Insights from male NMRI mice model.

BACKGROUND AND AIM: Chemotherapy, particularly with methotrexate (MTX), often elicits testicular toxicity, leading to impaired spermatogenesis and hormone imbalances. This study aimed to investigate the potential protective effects of selenium (Se) against MTX-induced testicular injury. MATERIALS AND METHODS: Male mice were divided into control, MTX, Se, and MTX + Se groups. Histopathological examination involved the preparation of testicular tissue sections using the Johnsen's tubular biopsy score (JTBS) for spermatogenesis evaluation. Biochemical tests included the assessment of testosterone, malondialdehyde (MDA), luteinizing hormone (LH), and follicle-stimulating hormone (FSH) levels. Real-time quantitative polymerase chain reaction (RT-qPCR) was employed to analyze the expression of caspase 3 (casp3), tumor protein 53 (p53), B-cell lymphoma 2 (Bcl2), and Bcl2-associated X protein (Bax) genes. Statistical analysis was performed using ANOVA and Tukey's tests (p < .05). RESULTS: Histopathological analysis revealed significant testicular damage in the MTX group, with decreased spermatogenesis and Leydig cell count, while Se administration mitigated these effects, preserving the structural integrity of the reproductive epithelium. Biochemical analysis demonstrated that MTX led to elevated malondialdehyde (MDA) levels and reduced testosterone, LH, and FSH levels, suggesting oxidative stress and Leydig cell dysfunction. Gene expression analysis indicated that MTX upregulated proapoptotic genes (casp3, p53, and bax) while downregulating the antiapoptotic Bcl2 gene. In contrast, Se treatment reversed these trends, highlighting its potential antiapoptotic properties. CONCLUSION: Our findings underscore the potential of Se as a therapeutic agent to mitigate the reproductive toxicity associated with MTX-induced testicular injury. Se exerts protective effects by regulating oxidative stress, preserving hormone balance, and modulating apoptotic pathways. These results suggest that Se supplementation could be a promising strategy to alleviate chemotherapy-induced testicular damage and preserve male fertility.

Engineered mesoporous silica nanoparticles, new insight nanoplatforms into effective cancer gene therapy.

The use of nucleic acid to control the expression of genes relevant to tumor progression is a key therapeutic approach in cancer research. Therapeutics based on nucleic acid provide novel concepts for untreatable targets. Nucleic acids as molecular medications must enter the target cell to be effective and obstacles in the systemic delivery of DNA or RNA limit their use in a clinical setting. The creation of nucleic acid delivery systems based on nanoparticles in order to circumvent biological constraints is advancing quickly. The ease of synthesis and surface modification, biocompatibility, biodegradability, cost-effectiveness and high loading capability of nucleic acids have prompted the use of mesoporous silica nanoparticles (MSNs) in gene therapy. The unique surface features of MSNs facilitate their design and decoration for high loading of nucleic acids, immune system evasion, cancer cell targeting, controlled cargo release, and endosomal escape. Reports have demonstrated successful therapeutic outcomes with the administration of a variety of engineered MSNs capable of delivering genes to tumor sites in laboratory animals. This comprehensive review of studies about siRNA, miRNA, shRNA, lncRNA and CRISPR/Cas9 delivery by MSNs reveals engineered MSNs as a safe and efficient system for gene transfer to cancer cells and cancer mouse models.

The effects of pomegranate peel extract on the gene expressions of antioxidant enzymes in a rat model of alloxan-induced diabetes.

This study was conducted to evaluate the anti-diabetic and antioxidant effects of hydroalcoholic pomegranate peel extract (APE) in alloxan-induced diabetes rat models. We divided 60 rats into the following six equal groups (n = 10): Healthy control; diabetic control (100 mg/kg alloxan); sham + glibenclamide (10 mg/kg); diabetic + glibenclamide (10 mg/kg); sham + APE (200 mg/kg) and diabetic + APE (200 mg/kg). After 8 weeks, kidneys were taken out for biochemical and molecular studies. Following APE treatment, biochemical parameters including malondialdehyde (MDA), and glutathione (GSH), glutathione peroxidase (GPx), catalase (CAT), superoxide dismutase (SOD) significantly induced in the treated group as compared with the control group (p < 0.05). Also, gene expression of GPx (3-fold), CAT (2.6-fold), and SOD (1.5-fold) were increased as compared to controls (p < 0.05). Overall, our results indicated that pomegranate can be used as an antioxidant agent to reduce complications from diseases associated with oxidative stress.

Effect of Cerium Oxide Nanoparticles on the Expression of Developmental and Apoptosis Genes of Testicular Tissue in 6-Day-Old NMRI Mice Fetuses.

Cerium oxide (CeO2) has potential applications in medicine and various consumer products. This study investigated the effect of CeO2 on the expression of genes associated with apoptosis and testicular development in mouse embryos. The experimental groups of pregnant mice were injected intraperitoneally with CeO2 at a concentration of 10 mg/kg on days 7 and 14 of pregnancy. Six days after birth, the testicles of neonatal male mice were collected for mRNA expression determination using real-time PCR, protein expression analysis by immunohistochemistry, and apoptotic cell population determination using the TUNEL assay. The results showed that the mRNA expression of the Bax, Caspase-3, and Gsk3-ß genes, unlike the Bcl2 gene, decreased significantly in the experimental group compared to the control group. The expression ratio of Bax/Bcl2 in the experimental group was lower than in the control group. A similar trend was observed in the population of apoptotic cells. In the experimental group, the expression levels of, Gata4, Sox8, and Rad54 at both the mRNA and protein levels increased significantly compared to the control group. Based on the results of this study, CeO2 at a concentration of 10 mg/kg, in addition to producing anti-apoptotic effects on the testicular cells of neonatal mice, can increase the expression of genes involved in testicular development and performance. The current experimental study proved the protective effects of 10 mg/kg CeO2 in developmental and apoptosis genes of testicular tissue in 6-day-old NMRI mice fetuses; however, more experiments are required to evaluate the possible side effects and interactions.