Exploring the combined anti-cancer effects of sodium butyrate and celastrol in glioblastoma cell lines: a novel therapeutic approach.

Glioblastoma, a highly aggressive and lethal brain cancer, lacks effective treatment options and has a poor prognosis. In our study, we explored the potential anti-cancer effects of sodium butyrate (SB) and celastrol (CEL) in two glioblastoma cell lines. SB, a histone deacetylase inhibitor, and CEL, derived from the tripterygium wilfordii plant, act as mTOR and proteasome inhibitors. Both can cross the blood-brain barrier, and they exhibit chemo- and radiosensitive properties in various cancer models. GB cell lines LN-405 and T98G were treated with SB and CEL. Cell viability was assessed by MTT assay and IC50 values were obtained. Gene expression of DNA repair, apoptosis, and autophagy-related genes was analyzed by RT-PCR. Cell cycle distribution was determined using flow cytometry. Viability assays using MTT assay revealed IC50 values of 26 mM and 22.7 mM for SB and 6.77 µM, and 9.11 µM for CEL in LN-405 and T98G cells, respectively. Furthermore, we examined the expression levels of DNA repair genes (MGMT, MLH-1, MSH-2, MSH-6), apoptosis genes (caspase-3, caspase-8, caspase-9), and an autophagy gene (ATG-6) using real-time polymerase chain reaction. Additionally, flow cytometry analysis revealed alterations in cell cycle distribution following treatment with SB, CEL and their combination. These findings indicate that SB and CEL may act through multiple mechanisms, including DNA repair inhibition, apoptosis induction, and autophagy modulation, to exert their anti-cancer effects in glioblastoma cells. This is the first study providing novel insights into the potential therapeutic effects of SB and CEL in glioblastoma.

Adipose delivered stem cells protect liver after ischemia-reperfusion injury by controlling autophagy.

OBJECTIVE: Ischemia-reperfusion(I/R) injury is an unavoidable side effect of liver surgery and transplantation. A potentially useful tool for cellular therapy and tissue engineering is adipose-derived stem cells (ADSCs).The process of autophagy is used by the cell to break down inappropriate molecules.The study's goal was to examine the impact of ADSCs on the autophagic pathway after rat hepatic ischemia-reperfusion injury. MATERIAL AND METHODS: Thirty male rats used in our study were divided into control, ADSC, ischemia, I/R, and I/R+ ADSC groups (n = 6). Liver tissues were stained with hematoxylin-eosin and histological changes were evaluated with Suzuki scoring. Immunoexpressions of transforming growth factor (TGF-ß) and autophagy markers LC3B, p62 were analyzed using the immunohistochemical method. RESULTS: As a result of histological evaluation the ischemia and I/R groups displayed sinusoid congestion, vacuolization, and necrosis in liver tissues. We showed that the immunostaining of LC3B and TGF- ß were elevated, and p62 decreased in the rat liver from ischemia and I/R groups when compared to the control group. CONCLUSION: ADSCs reduced the excessive level of autophagy and structural damage to hepatocytes and the pathological alterations in the liver after ischemia-reperfusion injury.

Evaluation of the effectiveness of oxytocin and enalapril in the prevention of epidural fibrosis developed after laminectomy in rats.

INTRODUCTION: Except for methylprednisolone, there is no current low-cost and low-side-effect drug/barrier method to prevent epidural fibrosis after spine surgery. However, the use of methylprednisolone has led to substantial controversy because of its serious side effects on wound healing. This study aimed to evaluate the effects of enalapril and oxytocin on preventing the development of epidural fibrosis in a rat laminectomy model. MATERIALS: Under sedation anesthesia, T9, T10, and T11 laminectomy was performed on 24 Wistar Albino male rats. The animals were then separated into four groups; Sham group (only laminectomy was performed; n = 6), MP group (laminectomy was performed and 10 mg/kg/day methylprednisolone was administered intraperitoneally (ip) for 14 days; n = 6), ELP group (laminectomy was performed and 0.75 mg/kg/day enalapril was administered ip for 14 days; n = 6), OXT group (laminectomy was performed and 160 µg/kg/day oxytocin was administered ip for 14 days; n = 6). Four weeks after the laminectomy, all the rats were euthanised, and the spines were removed for histopathological, immunohistochemical, and biochemical examinations. RESULTS: Histopathological examinations revealed that the degree of epidural fibrosis (X2=14.316, p = 0.003), collagen density (X2=16.050, p = 0.001), and fibroblast density (X2=17.500, p = 0.001) was higher in the Sham group and lower in the MP, ELP, and OXT groups. Immunohistochemical examinations showed that collagen type 1 immunoreactivity was higher in the Sham group and lower in the MP, ELP, and OXT groups (F = 54.950, p < 0.001). The highest level of α-smooth muscle actin immunoreactivity was seen in the Sham and OXT groups, and the lowest was in the MP and ELP groups (F = 33.357, p < 0.001). Biochemical analysis revealed that tissue levels of TNF-α, TGF-ß, IL-6, CTGF, caspase-3, p-AMPK, pmTOR, and mTOR/pmTOR were higher in the Sham group and lower in MP, ELP, and OXT groups (p < 0.05). The GSH/GSSG levels were lower in the Sham group and higher in the other three groups (X2=21.600, p < 0.001). CONCLUSION: The study results showed that enalapril and oxytocin, which are known to have anti-inflammatory, antioxidant, anti-apoptotic, and autophagy-related regenerative properties, could reduce the development of epidural fibrosis after laminectomy in rats.

The immunoexpression of valosin-containing protein and small VCP-interacting protein in rat ovaries.

Numerous cellular processes are controlled by the ubiquitin-proteasome-mediated degradation pathway, involve the 97-kDa valosin-containing protein (p97/VCP). Small p97/VCP-interacting protein (SVIP) was first discovered as one of the novel androgen-responsive genes as well as one of the many cofactors controlling p97/VCP. The aim of the study was to investigate localization and immunoexpression of p97/VCP and SVIP in rat ovarian tissue. The histomorphological examination of rat ovarian tissue was performed by using haematoxylin-eosin (HE) staining. Using the immunohistochemical technique, cellular location and expression of p97/VCP and SVIP in rat ovarian tissue were examined. The nuclear and cytoplasmic immunoexpression of p97/VCP and SVIP was observed in the different stages of ovarian follicles and corpus luteum in the rat ovaries. The immunolocalization of SVIP and VCP in the rat ovaries suggest that they may be involved in the oogenesis. Further studies should be performed about the function of the VCP and SVIP in the female reproductive tract.

Investigation of therapeutic effects of calcium dobesilate in cerebral hypoxia/ reperfusion injury in rats.

OBJECTIVES: Cerebral stroke is a serious clinical condition in which oxidative stress, inflammation, necrosis, apoptosis, and autophagy play important roles in its pathogenesis. This study investigated the neuroprotective and healing effects of calcium dobesilate (CD) on cerebral hypoxia/reperfusion injury in rats. METHODS: Forty Wistar albino male rats, each weighing 300-350 g, were separated into the Control group (no surgery and no pharmacological agent was administered); Sham-A group (only surgery was performed); DBL-A group (surgery was performed and CD 100 mg/kg/day was administered intraperitoneally for 3 days); Sham-C group (only surgery was performed); and DBL-C group (surgery was performed and 100 mg/kg/day CD was administered intraperitoneally for 10 days). Under sedation anesthesia, the bilateral common carotid arteries of all rats except the Control group were clipped for 30 min. After 4 h, the CD was given to the relevant groups, and then, all subjects were euthanized at scheduled times. The brain of each animal was removed for histopathological (hematoxylin and eosin staining), immunohistochemical (beclin-1, anti-MHC class II and anti-CD-68 staining), and biochemical (TNF, IL-1ß, IL-6, caspase-3, GSH/GSSG, malondialdehyde, protein carbonyl, LC3II/LC3I, and beclin-1 levels) evaluations. RESULTS: It was observed that CD could reduce necrosis and mitigate polarization of microglia to the M1 phenotype, autophagy, free oxygen radicals, protein carbonylation, lipid peroxidation, IL-1ß, IL6, TNF, caspase-3, beclin-1, and LC3II/LC3I levels in acute and chronic periods of hypoxia/reperfusion injury. CONCLUSION: From these results, it was observed that CD treatment could reduce neuronal necrosis and create anti-inflammatory, anti-edema, anti-oxidant, anti-apoptotic, and anti-autophagic effects in hypoxia/reperfusion injury in rats.

The protective effect of erythropoietin on ischemia- reperfusion injury caused by ovarian torsion-detorsion in the experimental rat model.

Ovarian torsion is one of the most dangerous gynecological emergencies requiring surgery. A total of 50%-90% ovarian torsion cases are caused by physiological cysts, endometriosis, and other benign or malignant ovarian neoplasms. The aim of the study was to investigate the effects of erythropoietin (EPO) treatment on ischemia/reperfusion (IR) injury caused by ovarian torsion/detorsion (T/D) injury. Thirty female Wistar albino rats were divided into five groups as follows: Group I: Control; Group II: Torsion (T); Group III: Torsion/Detorsion(T/D); Group IV: Torsion/Detorsion (T/D) + EPO; Group V: EPO. Sections of the ovaries were evaluated for histopathological changes with hematoxylin and eosin stain, a immunohistochemical assay for caspase 3 expression, and the TUNEL assay for apoptosis. Ovarian sections from torsion/detorsion and torsion groups showed more hemorrhage, vascular congestion, edema, degenerative granulosa, and stromal cells. Fewer histopathological changes were found in EPO and T/D + EPO groups. Caspase 3 and TUNEL positive cells were significantly increased in the torsion/detorsion group as compared with the other groups (p < 0.05). Treatment with erythropoietin decreased the number of caspase 3 and TUNEL positive cells. The results of the study showed that erythropoietin administration is effective for recovery from degenerative changes in the ovary induced by the torsion-detorsion injury.

Evaluation of the therapeutic effects of calcium dobesilate in sciatic nerve crush injury in rats.

INTRODUCTION: Proinflammatory cytokines released from nerve endings and surrounding injured tissue after nerve damage can prolong the inflammation process, delay nerve healing or result in poor quality nerve healing. In this case, due to the loss of function in the muscles innervated by the damaged nerve, the patient may have neurological and functional difficulties which may reduce the patient's quality of life and create an economic burden. Although the attempts of many pharmacological agents to heal crush injury of peripheral nerves have been recorded in literature, a drug that can provide adequate recovery of the crushed nerve and can be applied in daily life has not been defined as yet. This study aimed to assess the effects of calcium dobesilate on sciatic nerve crush injury in a rat model. METHODS: A total of 26 male Wistar albino rats were separated into four groups as follows: CONTROL group (healthy subjects, n=6); SHAM group (crush injury was created, n=6); MP group (after created crush injury, methylprednisolone was administered, n=7); and CAD group (after created crush injury, calcium dobesilate was administered, n=7). A crush injury was created, then the electrophysiological findings and sciatic nerve functional index (SFI) were recorded before euthanasia. After the euthanasia of all the rats, samples of the crushed nerve and gastrocnemius muscle were evaluated histopathologically, immunohistochemically, and biochemically. RESULTS: Both pharmacological agents were histopathologically effective in axon regeneration and repair. Calcium dobesilate did not preserve total muscle mass but was seen to prevent atrophy microscopically. Immunohistochemistry and biochemistry results showed that calcium dobesilate and methylprednisolone had anti-inflammatory, anti-oxidant, anti-apoptotic, and anti-autophagic activity in the crushed sciatic nerve. Neither calcium dobesilate nor methylprednisolone improved the nerve conductance level. SFI values obtained on day 30 from the CAD group were numerically closer to the values of the healthy animals but not at a statistically significant level. CONCLUSION: The study results demonstrated that calcium dobesilate could suppress inflammatory processes and provide histopathological and functional improvements in the injured nerve in rats. Therefore, further clinical studies are recommended to investigate in detail the therapeutic effects of calcium dobesilate on peripheral nerve crush injury.

Effects of regular whey protein consumption on rat thyroid functions

Background/aim: We aimed to investigate whether there was a significant difference in TSH, T3, T4 values and histopathologically evaluated thyroid tissues between rats that received isole hydrolyzed whey protein (IHWP) at different doses regularly and rats fed with only standard feed. Material & methods: Total 24 rats were randomly divided into three groups with 8 rats in each group. First group were fed with standard feed for 12 weeks. Second group were given standard feed + daily 0.3 g/kg IHWP and rats in the third group standard feed + 0.5 g/kg IHWP for 12 weeks. Blood samples were collected from all rats before and after IHWP administration. All rats were then sacrificed, and thyroid tissues were histopathologically examined. Results: Interfollicular connective tissue areas and TSH (0.35­4.90 µIU/L) were higher in the control group compared to 3 cc IHWP and 5 cc IHWP groups, while thyroid hormone T4 (0.7­1.48 ng/dL), and thyroid hormone synthesis parameters including intrafollicular colloid amount, follicular diameter, and epithelial height were significantly higher in 3 cc and 5 cc IHWP groups compared to the control. Conclusion: We think that regular daily use of IHWP may increase the synthesis of thyroid hormone due to its high amino acid content.

Investigation of the Effect of Alemtuzumab in an Experimental Spinal Cord Trauma Model in Rats.

OBJECTIVE: Spinal cord injuries generate the most negative response to medical treatment among all general body injuries. This important morbidity is thought to be caused by a complex secondary damage mechanism. In the present study, we examined the neuroprotective effects of alemtuzumab in a spinal cord trauma model. METHODS: We divided 24 Long-Evans male rats into 4 groups (n = 6 per group). Laminectomy was performed at T5-T8 in all groups. Trauma was applied using the Yasargil temporary aneurysm clip for 60 seconds at these spinal cord levels in all groups, except for group 1. Next, 1 mg/kg of alemtuzumab was administered to each rat in groups 3 and 4. A functional evaluation was performed on days 1, 3, and 5 in groups 1, 2, and 4, and the rats were then sacrificed. The rats in group 3 were sacrificed on the third postoperative day to observe the early effects of alemtuzumab. The biochemical examination findings of malondialdehyde and glutathione in plasma and tissue samples and histopathological findings of the spinal cord were evaluated and compared by statistical analysis. RESULTS: The inflammatory findings in the trauma group were not seen in either group treated with alemtuzumab. The clinical motor examination and inclined plane test results were also significantly better in these groups. CONCLUSION: Our results have shown that alemtuzumab might prevent spinal cord injury after trauma and is a histopathologically and biochemically strong anti-inflammatory, antioxidant, and neuroprotective agent.

The effect of antioxidants on angiogenesis in uterine transplantation.

The aim of this study was to investigate the effect of antioxidants on angiogenesis in uterine transplantation. We used 24 female rats equally divided into four groups: Group 1 had the uterus stored in HTK (Histidine-Tryptophan-Ketoglutarate) solution at 4 °C cold storage for 4 h. Group 2 had the uterine tissue stored in HTK solution combined with acetyl L-carnitine (10-8 M) for 4 h at +4 °C. The same procedures with Group 1 and 2 were repeated for 24 h for Groups 3 and 4, respectively. Histological investigation and immunohistochemical analysis were performed. Histological findings showed that storing donor uterus in HTK solution at +4° C for 24 h results in histological alteration in uterus. We also found that immunoreactivity of VEGFR-2 in all layers of rat uterus in Group 2 was lower than that in Group 1, and the expression of the uterus in Group 4 was lower than that in Group 3. We concluded that antioxidant acetyl L-carnitine, which was added to the organ preservation solution HTK, had prevented the formation of free radicals, and thus protected the uterus that was stored in short and long cold storage periods. Impact statement What is already known on this subject? Ischemia-reperfusion is a complex pathophysiological process involve in hypoxia and/or reoxygenation, ionic imbalance-induced oedema and acidosis, oxidative stress, mitochondrial uncoupling, coagulation and endothelium activation. The composition of preservation solutions must be adapted to the severity of ischaemia-reperfusion injuries to reduce cellular damage and inflammation and preserve graft functionality and integrity, thus improving short-term and long-term graft outcome. Clinicians use three types of composition of solution for static cold preservation: intracellular, intermediate and extracellular. HTK will be used frequently, especially with the consideration of lower price and more easy handling aspects. L-carnitine acts as an antioxidant, protects against free radicals and prevents mitochondrial damage. VEGFR-2 plays an important role in angiogenesis, chemotaxis, proliferation and migration of endothelial cells. What this study adds? In this study, we investigate the effect of antioxidants on angiogenesis in uterus transplantation. Our results showed that antioxidant acetyl L-carnitine that added to the organ preservation solution HTK, has prevented the formation of free radicals, thus protect the uterus that was stored in short and long cold storage periods. What the implications are for future studies? Therefore, we will contribute to the literature with the results of this study.

Possible effects of melatonin against rat uterus exposure to bisphenol A during neonatal period.

The aim of this study was to investigate the possible effects of melatonin on rat uterine tissue against exposure with bisphenol A (BPA) in the neonatal period. Twenty-four female rats were divided into four groups, (n=6) per group. Group I was used as a control (sesame oil + ethanol), group II was injected daily with (100 mg/kg) BPA by subcutaneously (sc) daily postnatal days (PND 0-10), group III was injected daily with (10 mg/kg) melatonin by sc for 10 days (PND 20-30), and group IV was injected daily with (100 mg/kg) BPA (PND 0-10) and (10 mg/kg) melatonin (PND 20-30). All rats were sacrificed in the same day of metestrus cycle, approximately PND 70. Histological analyses, immunostaining of B cell lymphoma 2 (Bcl-2), and cytochrome c and TUNEL assays were performed. According to our results, neonatal exposure to BPA accelerates onset of puberty, causes degenerative and morphometric changes on rat uterus, and increases apoptotic reaction rates. The immunoreactivity of Bcl-2 was decreased after BPA administration. In addition, immunoreactivity of Bcl-2 showed an increase after melatonin treatment. However, cytochrome c immunoreactivity was decreased after melatonin administration. Our results suggest that melatonin may have positive effects against BPA-induced degenerative changes on rat uterus.