Lipocalin-2 Deficiency Diminishes Canonical NLRP3 Inflammasome Formation and IL-1ß Production in the Subacute Phase of Spinal Cord Injury.

Spinal cord injury (SCI) results in the production of proinflammatory cytokines due to inflammasome activation. Lipocalin 2 (LCN2) is a small secretory glycoprotein upregulated by toll-like receptor (TLR) signaling in various cells and tissues. LCN2 secretion is induced by infection, injury, and metabolic disorders. In contrast, LCN2 has been implicated as an anti-inflammatory regulator. However, the role of LCN2 in inflammasome activation during SCI remains unknown. This study examined the role of Lcn2 deficiency in the NLRP3 inflammasome-dependent neuroinflammation in SCI. Lcn2-/- and wild-type (WT) mice were subjected to SCI, and locomotor function, formation of the inflammasome complex, and neuroinflammation were assessed. Our findings demonstrated that significant activation of the HMGB1/PYCARD/caspase-1 inflammatory axis was accompanied by the overexpression of LCN2 7 days after SCI in WT mice. This signal transduction results in the cleaving of the pyroptosis-inducing protein gasdermin D (GSDMD) and the maturation of the proinflammatory cytokine IL-1ß. Furthermore, Lcn2-/- mice showed considerable downregulation in the HMGB1/NLRP3/PYCARD/caspase-1 axis, IL-1ß production, pore formation, and improved locomotor function compared with WT. Our data suggest that LCN2 may play a role as a putative molecule for the induction of inflammasome-related neuroinflammation in SCI.

Combination of two miRNAs has a stronger effect on stimulating apoptosis, inhibiting cell growth, and increasing erlotinib sensitivity relative to single miRNA in A549 lung cancer cells.

Despite the dramatic efficacy of EGFR-TKIs, most of non-small cell lung cancer patients ultimately develop resistance to these agents. In this study, we explored the effects of miRNA-125a-5p and miRNA-145, alone or in combination, EGFR expression, cell growth and sensitivity of the NSCLC cells to erlotinib. The expression of EGFR was measured using RT-qPCR and Western blotting. The effect of miRNAs and erlotinib on cell growth and survival was assessed by trypan blue assay and MTT assay, respectively. Apoptosis was measured using ELISA cell death assay. We found that transfection of miRNA-125a-5p and miRNA-145 significantly inhibited the expression of EGFR mRNA and protein in a time-dependent manner (p < 0.05 vs. blank control or negative control miRNA). ANOVA and Bonferroni's test were used to ascertain significant differences between groups. Other experiments indicated that upregulation of each of miRNA-125a-5p or miRNA-145 inhibited cell growth, induced apoptosis, and markedly decreased the IC50 value of erlotinib in A549 lung cancer cells (p < 0.05). Moreover, the combination of two miRNAs showed a stronger effect on cells survival, apoptosis, and drug sensitivity, relative to single miRNA (p < 0.05). The results of our study indicate that the therapeutic delivery of miRNA-145 and miRNA-125a-5p to lung cancer may inhibit cell proliferation, trigger apoptosis, and sensitize lung cancer cells to EGFR-TKIs.

The effect of curcumin on NRF2/Keap1 signalling pathway in the epididymis of mouse experimental cryptorchidism.

Nrf2/Keap1 pathway, which prevents cellular damage against reactive oxygen species production, is disrupted in epididymis following cryptorchidism. In this study, we aimed to use curcumin (Cur) as an activator of Nrf2 to decrease the effects of disruption in this pathway caused by cryptorchidism. In this study, animals were randomly divided into following groups: control, sham-surgery, sham-vehicle, sham-Cur50, sham-Cur100 , cryptorchidism, cryptorchidism-vehicle, cryptorchidism-Cur50 and cryptorchidism-Cur100 . For cryptorchidism induction, the left testicle was removed from the scrotum and sutured to the abdominal wall. Two weeks after surgery, Cur was given orally to animals. After 1 month, sperm parameters and testis histopathology were analysed. The expression of Nrf2, NQO1, HO1, and Keap1 genes was evaluated by real-time polymerase chain reaction. Our data showed that Cur, especially at high doses, could improve sperm parameters and testis histopathology, which were damaged following cryptorchidism induction. The expression of HO1, NQO1, and Nrf2 genes, which had decreased in the cryptorchidism group, showed a significant increase after administration of Cur in a dose-dependent manner. Cur, by inducing the expression of genes involved in the Nrf2/Keap1 pathway, could reduce the adverse effects of cryptorchidism and might be used as adjuvant therapy for decreasing cryptorchidism complications before surgery.

Effect of bromelain on mast cell numbers and degranulation in diabetic rat wound healing.

OBJECTIVE: Many studies have revealed the prominent roles of mast cells in wound healing, including inflammatory reactions, angiogenesis and extracellular matrix reabsorption. In the present study, we aimed to assess the probable therapeutic features of bromelain on wound contraction and mast cell degranulation in wound healing in experimental diabetic animals. METHOD: Male rats were grouped as control, vehicle and experiment. Skin wounds were generated in all groups. Treatments were applied with distilled water and with bromelain (BR) intraperitoneally in the vehicle and experimental groups, respectively. Following skin wound generation, animals were euthanised on days 3, 5, 7 and 15. We gathered 16,800 microscopic images to count the mast cells and degranulation level (Image J software). The wound contraction index was assessed both microscopically (Image J software) and macroscopically (time-lapse photography). The meshwork evaluation method was used to assess wound healing. RESULTS: Time-lapse photography revealed that the BR significantly (p<0.05) accelerated wound contraction and healing. BR significantly (p<0.05) increased the total number of mast cells in all experimental groups on days 5 and 7. The count of grade III (degranulated) mast cells was reduced significantly (p<0.05) on days 5 and 7 in experimental groups compared to control and vehicle groups. CONCLUSION: In this study, the rate of wound healing was accelerated considerably following BR administration. In addition, this agent decreased the count of degranulated mast cells, leading to wound contraction and healing.

The effects of different training modalities on monocarboxylate transporters MCT1 and MCT4, hypoxia inducible factor-1α (HIF-1α), and PGC-1α gene expression in rat skeletal muscles.

The research literature suggests that different training modalities cause various patterns in training-induced genes expression. This study aimed to investigate the effects of moderate intensity continuous training (MICT) and isocaloric high intensity interval training (HIIT) on gene expression of monocarboxylate transporter 1 (MCT1) and 4 (MCT4), Peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α), and hypoxia inducible factor-1α (HIF-1α) in soleus and extensor digitorum longus (EDL) skeletal muscles of rats. Thirty male Sprague-Dawley rats were divided into 3 groups of control, MICT, and HIIT. Training protocols were performed according to the principle of overload for 8 weeks and 5 sessions per week. Then, the soleus and EDL muscles were extracted and the expression levels were analyzed using the real time PCR method. In the MICT group, only the EDL HIF-1α mRNA level was significantly higher than that of the control group (p < 0.05). In the HIIT group, however, mRNA levels of MCT4, PGC-1α, and HIF-1α in both muscles were significantly higher than those of the control group (p < 0.05). The comparison between the two training methods demonstrated that the gene expression levels of soleus and EDL MCT4, soleus PGC-1α, and soleus HIF-1α were significantly higher in the HIIT group compared to the MICT group (p < 0.05). There were also significant positive correlations between all mRNA levels of HIF-1α and corresponding mRNA levels of MCT4 (p < 0.05). HIIT caused greater positive responses in the gene expression of MCT4, PGC-1α, and HIF-1α compared to MICT.

Knockdown of Myeloid Cell Leukemia-1 by MicroRNA-101 Increases Sensitivity of A549 Lung Cancer Cells to Etoposide.

Background: Studies have shown that myeloid cell leukemia-1 (Mcl-1) is the target gene for microRNA -101 (miRNA-101), and decreased levels of miRNA-101 are associated with elevated levels of Mcl-1 and lung cancer survival. The objective of the present study was to investigate the effect of miRNA-101 on the sensitivity of A549 lung cancer cells to etoposide. Methods: The study was conducted during 2018 and 2019 at Arak University of Medical Sciences, Arak, Iran. The effect of miRNA-101 on Mcl-1 expression was assessed using reverse transcription-quantitative polymerase chain reaction 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT), and trypan blue exclusion assays were performed to determine the effect of treatments on cell survival and proliferation, respectively. The interaction between miRNA-101 and etoposide was evaluated using the combination index analysis of Chou-Talalay. Apoptosis was quantified using ELISA cell death assay. ANOVA and Bonferroni's tests were used to determine statistical differences between the groups (P<0.05). GraphPad Prism software (version 6.01) was used for data analysis. Results: The results showed that miRNA-101 clearly inhibited the expression of Mcl-1 and reduced the growth of A549 cells, relative to blank control and negative control miRNA (P<0.05). Transfection of miRNA-101 synergistically enhanced the sensitivity of the A549 cells to etoposide. Apoptosis assay data also showed that miRNA-101 triggered apoptosis and augmented the etoposide-mediated apoptosis. Conclusion: Up-regulation of miRNA-101 inhibited cell survival and proliferation, and sensitized A549 cells to etoposide by suppressing Mcl-1 expression. miRNA-101 replacement therapy can be considered as an effective therapeutic strategy in non-small cell lung cancer.

Protective effect of vitamin E on sperm parameters in rats infected with Candida albicans.

Candida albicans is one of the most frequent pathogens present in the reproductive system. The negative in vitro effects of C. albicans on sperm functions have previously been studied. The current study was undertaken to investigate the effects of C. albicans infection in vivo on sperm quality and to evaluate the efficacy of vitamin E administration in rats infected with C. albicans. In this study, 5 days after infection induction, animals were treated with vitamin E for 5 weeks. Thereafter, sperm parameters, lipid peroxidation (LPO), total antioxidant capacity (TAC), hormonal analysis and testis histology were evaluated. Based on the results, sperm parameters and TAC significantly reduced, while LPO and tissue damage increased (p ≤ .05) following the infection. Hormone analysis showed low LH and testosterone levels in serum of the infected rats. Treatment with vitamin E significantly (p ≤ .05) improved sperm quality and testis histology, increased TAC and reduced LPO. In addition, vitamin E administration significantly increased (p ≤ .05) serum LH and testosterone levels. These results clearly indicate that vitamin E is effective in attenuating the adverse effects of C. albicans infection on male fertility and could be used as a complementary treatment for patients who suffer from fertility disorders following C. albicans infection.

Effect of selenium and pentoxifylline on expression of CATSPER1 and 2 genes and FSH/LH levels in treated mice by dexamethasone.

Dexamethasone has deleterious effects on male fertility and sperm parameters. In this study, the effect of dexamethasone on expression of CATSPER1 and 2 genes was investigated. These two genes play an important role in sperm motility. Selenium and pentoxifylline were subsequently used to protect testis tissue against the destructive effects of dexamethasone. Each group received one of the following treatments for 7 days: dexamethasone (7 mg/kg), pentoxifylline (200 mg/kg), selenium (0.3 mg/kg), dexamethasone + pentoxifylline or selenium + dexamethasone. Animals in the control group received a normal saline injection. The expression of CATSPER1 and 2 genes was analysed by real-time PCR and serum levels of FSH and LH were determined with the enzyme-linked immunosorbent assay method. Based on the results, dexamethasone decreases not only CATSPER1 and 2 gene expression but also serum levels of LH (p ≤ 0.05); however, it has no effect on FSH (p > 0.05). Treating with selenium significantly increased the gene expression of both CATSPER1 and 2 (p ≤ 0.05), while pentoxifylline enhanced only CATSPER2 gene expression (p ≤ 0.05). These two antioxidants were shown to increase serum levels of LH (p ≤ 0.05). Our data suggest that selenium is more effective than pentoxifylline in overcoming adverse effects of dexamethasone on male fertility.

Effect of Omegaven on mast cell concentration in diabetic wound healing.

Diabetic wound healing is a complicated process. In all over the world 15% of 200 million diabetic people suffer from diabetic foot problems. Mast cells are known to participate in three phases of wound healing: the inflammatory reaction, angiogenesis and extracellular-matrix reabsorption. The inflammatory reaction is mediated by released histamine and arachidonic acid metabolites. Omega-3 fatty acids alter proinflammatory cytokine production during wound healing which affects the presence of inflammatory cells in wound area as well, but how this events specifically influences the presence of mast cells in wound healing is not clearly understood. This study is conducted to determine the effect of Omegaven, eicosapentaenoic (EPA) and docosahexaenoic (DHA) on pattern of presence of mast cells in diabetic wound area. Diabetic male wistar rats were euthanized at 1, 3, 5, 7 and 15 days after the excision was made. To estimate the number of mast cells histological sections were provided from wound area and stained with toluidine blue. In this relation wound area (8400 microscopic field, 45.69 mm2) were examined by stereological methods by light microscope. We found that comparing experimental and control group, omega-3 fatty acids significantly decreased wound area in day 7 and also the number of grade three mast cells in day 3 and 5. We also found that wound strength has significantly increased in experimental group at day 15.

High-level expression and purification of soluble bioactive recombinant human heparin-binding epidermal growth factor in Escherichia coli.

Heparin-binding epidermal growth factor (HB-EGF) is a member of highly conserved superfamily of proteins that has potential mitogenic activity and stimulates differentiation and migration of various cell types. Since HB-EGF has three intra-molecular disulfide bonds, a high expression pattern of active HB-EGF in an E. coli expression system was not successfully established. The aim of this study was to increase production of soluble bioactive recombinant human HB-EGF in E. coli by modifying growth conditions and codon optimization. The open reading frame codons of human HB-EGF were optimized to achieve high level expression in E. coli. The optimized codon was amplified, cloned into plasmid pET-32a, and transformed into E. coli BL21 for further expression. The cultivation parameters (temperature and inducer) were optimized to produce a high yield of soluble HB-EGF. The fusion protein was purified by Nickel-nitrilotriacetic acid (Ni-NTA) affinity chromatography. Amethylthiazole tetrazolium assay was used to evaluate the bioactivity of the produced recombinant protein. After codon optimization, the codon adaptation index (CAI) was increased from 0.255 in native gene to 0.829 using the optimized sequence. By lowering the temperature to 22°C and the inducer to 0.4 µM, we obtained 35% soluble expression of recombinant and biologically active human HB-EGF. Our data demonstrate that codon optimization increases the yield of HB-EGF in an E. coli expression system. Furthermore, the chosen modifications in cell culturing increase the solubility of recombinant human HB-EGF.

The antiapoptotic effects of conditioned medium from bone marrow-derived mesenchymal stromal stem cells on cyclophosphamide-induced testicular damage in rat: An experimental study.

Background: Cyclophosphamide (CP) has some negative effects on the reproductive system. Stem cells and their metabolites are being utilized to enhance fertility after chemotherapy. Objective: This study aimed to investigate the impact of conditioned medium (CM) derived from bone marrow mesenchymal stromal stem cells (BM-MSCs) on the toxic effects of CP on testicles. Materials and Methods: BM-MSCs were isolated, a CM was collected and 25-fold concentrated. 24 male Wistar rats (8 wk, 200-250 gr) were randomly divided into following groups: control, CP, CP+ Dulbecco's Modified Eagle Medium (DMEM), CP+CM. CP was given at a single dose of 100 mg/kg. 2 wk after the CP administration, CM was injected into the testicular efferent duct. Sperm parameters, testicular histopathology, and the level of testosterone were analyzed 2 months after treatment. The expression of B-cell lymphoma 2 (Bcl2) and Bcl2-associated X protein (Bax) genes were evaluated by real-time polymerase chain reaction. Results: CP had a negative effect on testis histology (p < 0.001) and sperm quality (p < 0.001). It changed the expression of genes associated with apoptosis (p < 0.001). Treatment with CM reduced the expression of Bax (p < 0.001), while significantly increasing the expression of Bcl2 (p = 0.01). It improved sperm count (p = 0.03), viability (p < 0.001), motility (p < 0.001), spermatogonial count (p < 0.001), and epithelial thickness of testicular tubules (p = 0.02). Conclusion: These findings suggest that CM produced from BM-MSCs may be valuable for therapeutic approaches in reproductive medicine and may lessen the side effects of CP.

Protective effects of Withania somnifera against cyclophosphamide-induced testicular damage in rats.

Objective: Cyclophosphamide (CP) is an alkylating agent commonly used in cancer treatment. It is known to have detrimental effects on the reproductive system, including the potential to cause infertility. Recently, herbal remedies have gained traction as a complementary approach to addressing these side effects. In this study, our goal was to investigate whether the aqueous-alcoholic extract of Withania somnifera (WS) could mitigate the adverse impacts of CP on testicular tissue. Methods: Animals were randomly assigned to one of the following groups: control, WS (500 mg/kg), CP (100 mg/kg), CP+WS pre-treatment, and CP+WS post-treatment. WS was administered orally through gavage for 1 month. We assessed sperm parameters, testicular histopathology, and the expression of the Bax and Bcl2 genes in the experimental groups. Results: Sperm parameters (including count, viability, and motility), the number of spermatogonia, the seminiferous tubule diameter, and Bcl2 gene expression, significantly decreased after CP injection (p<0.05). Conversely, the number of immotile sperm and Bax gene expression significantly increased (p<0.05). Treatment with WS, especially when administered as a pre-treatment, ameliorated the sperm parameters, histological alterations, and the expression of apoptosis-related genes (p<0.05). Conclusion: The data suggest that WS may mitigate the detrimental effects of CP on testicular tissue by reducing apoptosis. Consequently, WS has the potential to be used as an adjunctive therapy to reduce the complications associated with CP treatment.

Resveratrol ameliorates spermatogenesis by increasing protamine 1, 2 and HSPA2 expression in experimental varicocele rat model.

OBJECTIVES: Varicocele is a common cause of male infertility associated with an elevated testicular temperature that induces apoptosis, spermatogenesis dysfunction, and affects sperm parameters. In this study, we investigate the probable therapeutic effects of resveratrol (RES), a natural phytoalexin, against varicocele. MATERIALS AND METHODS: In this study, 48 male Wistar rats randomly divided into 8 groups: normal, sham, normal+RES (20 and 50mg/kg), varicocele, varicocele+ethanol and varicocele+RES (20 and 50mg/kg). Incomplete closure of the left renal vein was used for varicocele induction and two months later, RES was administrated orally for 60 days. Then, sperm parameters, DNA fragmentations, chromatin density, and testis histopathology were analyzed. In addition, HSPA2, protamine 1, and 2 expression levels were evaluated using real-time PCR. RESULTS: According to our results, resveratrol treatment improved sperm parameters, testis histopathology, DNA fragmentation, and chromatin maturation which damaged follow varicocele (p≤0.05). Also, it increased HSPA2, protamine 1, and 2 expression levels significantly in both doses (p≤0.05). CONCLUSION: Resveratrol potentially attenuates varicocele-induced spermatogenic impairments by its antioxidant features and regulates spermatogenic gene expression undergoing DNA fragmentation, so leads histopathological properties of tissues to physiological parameters.

The time course of nod-like receptor family pyrin domain containing 3 inflammasome complex expressions in the testis tissue of an experimental varicocele rat model: An experimental study.

Background: Varicocele is characterized by abnormal dilation of the testicular vein, which results in hypoxia, the accumulation of reactive oxygen species, and the production of proinflammatory cytokines. It seems that a group of cytosolic receptors named nod-like receptor family, pyrin domain containing 3 (NLRP3) inflammasome, is activated and involved in the pathogenesis of varicocele. Objective: We aim to determine the time course of NLRP3 inflammasome expression in the testis tissue following varicocele induction. Materials and Methods: In this experimental study, 36 adult Wistar rats (8 wk, 200-250 gr) were used. For the varicocele induction, the left renal vein was partially ligated. The mRNA levels of NLRP3, apoptosis-associated speck-like protein containing a caspase recruitment domain, and caspase-1 were evaluated by real-time polymerase chain reaction at 1, 2, 4, 8, and 12 wk after varicocele induction. Results: Results showed that the gene expression of NLRP3 inflammasome component including NLRP3, apoptosis-associated speck-like protein containing a caspase recruitment domain, and caspase-1 did not alter during week 1, 2, 4, and 8 after operation (p = 0.09). 12 wk after varicocele induction, gene expression levels were significantly up-regulated (p = 0.02). Conclusion: Our data provides clear evidence that varicocele stimulates inflammasome activation in the testis tissue 12 wk after the operation, and this time is required for investigating NLRP3 activity in the varicocele rat model.

Regulation of Inflammasomes by Application of Omega-3 Polyunsaturated Fatty Acids in a Spinal Cord Injury Model.

Omega-3 polyunsaturated fatty acids (PUFA n3) ameliorate inflammation in different diseases and potentially improve neurological function after neuronal injury. Following spinal cord injury (SCI), inflammatory events result in caspase-1 mediated activation of interleukin-1 beta (IL-1b) and 18. We aim to evaluate the neuroprotective potency of PUFA n3 in suppressing the formation and activation of inflammasomes following SCI. Male Wistar rats were divided into four groups: control, SCI, SCI+PUFA n3, and SCI+Lipofundin MCT (medium-chain triglyceride; vehicle). PUFA n3 or vehicle was intravenously administered immediately after SCI and every 24 h for the next three days. We analyzed the expression of NLRP3, NLRP1, ASC, caspase-1, IL-1b, and 18 in the spinal cord. The distribution of microglia, oligodendrocytes, and astrocytes was assessed by immunohistochemistry analysis. Behavioral testing showed significantly improved locomotor recovery in PUFA n3-treated animals and the SCI-induced upregulation of inflammasome components was reduced. Histopathological evaluation confirmed the suppression of microgliosis, increased numbers of oligodendrocytes, and the prevention of demyelination by PUFA n3. Our data support the neuroprotective role of PUFA n3 by targeting the NLRP3 inflammasome. These findings provide evidence that PUFA n3 has therapeutic effects which potentially attenuate neuronal damage in SCI and possibly also in other neuronal injuries.

MiRNA-Mediated Knock-Down of Bcl-2 and Mcl-1 Increases Fludarabine-Sensitivity in CLL-CII Cells.

BACKGROUND: Over-expression of anti-apoptotic proteins such as Bcl-2 and Mcl-1 is associated with resistance to chemotherapeutic agents such as fludarabine. Moreover, an inverse relationship between miRNA-15a levels with Bcl-2 and Mcl-1 expression has been observed in CLL patients. In this study, the effect of miRNA-15a on apoptosis and sensitivity of the CLL cells to fludarabine was investigated. METHODS: After treatments, the Mcl-1 and Bcl-2 expression levels were quantified by RT-qPCR. Trypan blue assay was used to explore the effects of miRNA-15a and fludarabine on cell proliferation. The cytotoxicity was measured using MTT assay and combination index analysis. Cell death was determined using cell death detection ELISA assay and caspase-3 activity assay Kits. RESULTS: Results showed that miRNA-15a clearly decreased the mRNA levels of Bcl-2 and Mcl-1 in a time dependent manner, which led to CLL-II cell proliferation inhibition and enhancement of apoptosis (p < 0.05, relative to control). Transfection of the miRNA-15a synergistically reduced the cell survival rate and lowered the IC50 value of fludarabine. Furthermore, miRNA-15a significantly enhanced the apoptotic effect of fludarabine. CONCLUSIONS: Our data propose that suppression of Bcl-2 and Mcl-1 by miRNA-15a can effectively inhibit the cell proliferation and sensitize CLL cells to fludarabine. Therefore, miRNA-15a can be considered as a potential therapeutic target in CLL resistant patients.

MiRNA-7 Replacement Effect on Proliferation and Tarceva-Sensitivity in U373-MG Cell Line.

BACKGROUND: Deregulation of the EGFR signaling pathway activity has been shown to can be effective in resistance to EGFR-TKIs, such as Tarceva (erlotinib), in glioblastoma cells. In addition, reports have shown that the reduction of miRNA-7 expression levels is associated with an increase in the expression of EGFR. Here, we evaluated the effect of miRNA-7 on EGFR expression and sensitivity of the U373-MG glioblastoma to erlotinib. METHODS: The effect of miRNA-7 on EGFR expression was examined using RT-qPCR and western blotting. Trypan blue and MTT assays were performed to explore the effect of treatments on cell growth and survival, respectively. The combination index analysis was used to evaluate the interaction between drugs. Apoptosis was measured by ELISA cell death assay. RESULTS: We showed that miRNA-7 markedly inhibited the expression of EGFR and decreased the growth of glioblastoma cells, relative to blank control and negative control miRNA (p < 0.05). Introduction of miRNA-7 synergistically increased the sensitivity of the U373-MG cells to erlotinib. Results of apoptosis assay demonstrated that miRNA-7 can trigger apoptosis and enhance the erlotinib-mediated apoptosis. CONCLUSIONS: Our results show that miRNA-7 plays a critical role in the growth, survival and sensitivity of the U373-MG cells to erlotinib by targeting EGFR. Thus, miRNA-7 replacement therapy can become an effective therapeutic procedure in glioblastoma.

Presence of The NLRP3 Inflammasome Components in Semen of Varicocele Patients.

BACKGROUND: Varicocele is a common cause of male infertility with multifactorial etiology. Inflammation is a characteristic pathological event that occurs in the testis tissue following the varicocele. The aim of this study was to investigate expression of nod-like receptor family, pyrin domain containing 3 (NLRP3) inflammasome components and cytokines in semen of varicocele and control subjects. MATERIALS AND METHODS: In this case-control study, seminal plasma was collected from 32 varicocele patients (with grades 2 and 3) and 20 fertile men as control group. Semen analysis was performed in all subjects. Concentrations of interleukin-1b (IL-1b), IL-18 and caspase-1 in seminal plasma were measured by enzyme-linked immunosorbent assay (ELISA). Apoptosis-associated speck-like protein containing a caspase activation and recruitment domain, in addition to NALP3 were identified in seminal plasma by Western blot. Statistical significance between the mean values was determined by student's t test. RESULTS: According to our data, the level of IL-1b was significantly (P=0.03) increased in the seminal plasma of varicocele patients, compared to the control subjects. We analyzed amount of IL-18 in the both groups. The level of this interleukin was markedly (P=0.002) decreased in varicocele patients. No change was observed in the level of caspase-1 in both groups. Western blot analysis revealed that apoptosis associated speck-like protein (ASC, P=0.0002) and NLRP3 (P=0.005) were significantly elevated in the semen of varicocele patients. CONCLUSION: This study provides the first evidence of activation of NLRP3 components in semen of men with varicocele.

Targeting Epidermal Growth Factor Receptor by MiRNA-145 Inhibits Cell Growth and Sensitizes NSCLC Cells to Erlotinib.

Background: Despite effective activity of epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs), such as erlotinib, all non-small cell lung cancer (NSCLC) patients eventually acquire resistance to these agents. Studies have demonstrated that down-regulation of miRNA-145 leads to enhancement of EGFR expression, cell proliferation and metastasis. The aim of this study was to investigate the effect of miRNA-145 on sensitivity of the A549 NSCLC cells to erlotinib. Methods: Quantitative real-time PCR was used to examine the effect of miRNA-145 on EGFR expression. The effect of miRNA-145 on cell growth and sensitivity the lung cancer cells to erlotinib was examined by trypan blue and MTT assays, respectively. The combination index was calculated using the non-constant method of Chou-Talalay. Apoptosis was determined by ELISA cell death assay. Results: We found that miRNA-145 was markedly suppressed the expression of EGFR and inhibited the cancer cell growth, relative to blank control and negative control miRNA (p<0.05). Pretreatment with miRNA-145 synergistically enhanced the sensitivity of the lung cancer cells to erlotinib. Results of apoptosis assay revealed that miRNA-145 can induce apoptosis and increase the erlotinib-mediated apoptosis. Conclusions: Our data demonstrate that miRNA-145 play a critical role in the lung cancer cell growth, survival and EGFR-TKIs resistance possibly by regulation of EGFR. Therefore, miRNA-145 replacement therapy can become a new therapeutic strategy in lung cancer.

Possible antioxidant mechanism of coenzyme Q10 in diabetes: impact on Sirt1/Nrf2 signaling pathways.

Oxidative stress is a major complication in diabetes mellitus. The aim of this study was to investigate potential antioxidant activity of coenzyme Q10 (Co Q10) against hyperglycemia-induced oxidative stress in diabetic rat and unraveling its mechanism of action by focusing on silent information regulator 1 (Sirt1) and nuclear factor E2-related factor 2 (Nrf2) mRNA expression level. Furthermore, the activity of two Nrf2-dependent antioxidant enzymes (superoxide dismutase and catalase) in the liver of diabetic rats was studied. After induction of diabetes in rats using streptozotocin (55 mg/kg), rats were divided into five groups of six each. Groups 1 and 2 (healthy control groups) were injected with isotonic saline or sesame oil; group 3 received Co Q10 (10 mg /Kg /day), group 4, as a diabetic control, received sesame oil; and group 5 was diabetic rats treated with Co Q10. Afterwards, serum and liver samples were collected, and oxidative stress markers, lipid profile, as well as the expression of Sirt1 and Nrf2 genes were measured. Diabetes induction significantly reduced expression level of Sirt1 and Nrf2 mRNAs and also declined catalase, superoxide dismutase activities, and total thiol groups levels in diabetic group in comparison to healthy controls, while a significant increase was found in the levels of malondialdehyde and lipid profile. Co Q10 treatment significantly up-regulated Sirt1 and Nrf2 mRNA levels along with an increase in catalase activity in diabetic group as compared with untreated diabetic rats. Furthermore, Co Q10 caused a marked decrease in malondialdehyde levels and significantly improved lipid profile. Our data demonstrated that Co Q10 may exert its antioxidant activity in diabetes through the induction of Sirt1/Nrf2 gene expression.