Biotechnic & Histochemistry最新文献

Testicular torsion, a medical condition contributing to male infertility, results in severe scrotal pain and ischemia. Oxidative stress factors contribute to germ cell death following surgical reperfusion, suggesting postoperative pharmacotherapy could mitigate testicular ischemia/reperfusion (I/R) injury. Ivermectin, a GABA receptor modulator for treating parasitic infections such as onchocerciasis, has demonstrated anti-oxidative stress and anti-apoptotic properties. Therefore, this study aimed to evaluate the efficacy of ivermectin administration after detorsion using a rat model of testicular torsion/detorsion (T/D). This study utilized 40 male Wistar rats weighing 200-250 grams. The studied groups were the sham (no T/D induction), control (T/D and no treatment), and T/D following administration of ivermectin administration and the doses of 1, 2, and 5 mg/kg intraperitoneally. For I/R surgery, both testes were twisted 720 degrees clockwise for 4 h to cause torsion. Subsequently, the levels of malondialdehyde (MDA) and superoxide dismutase (SOD) activity were assessed using ELISA, while the expression of GABA_B receptors and apoptosis were examined using the immunohistochemical method. Histopathological alterations were evaluated with hematoxylin and eosin (H&E) staining to evaluate cell counts and seminiferous tubule diameters. Administration of ivermectin effectively reduced oxidative stress levels, as evidenced by decreased MDA levels and increased SOD activity, compared to the control group (P < 0.01 and P < 0.05, respectively). Ivermectin also modulated the expression of elevated GABA_B receptors and caspase 3 (P < 0.05 and P < 0.001, respectively). Furthermore, histopathological analysis revealed that ivermectin prevented germ cell degeneration, edema, and hemorrhage in the testis (P < 0.05). Ivermectin may be a potential treatment option for protecting against testicular torsion and may have beneficial effects on mitigating germ cell death, oxidative stress, and GABA_B receptor modulation. Further research is needed to explore dosages, long-term impacts, and clinical trials to determine the potential for patients with testicular torsion.

Amyloidosis is caused by the extracellular deposition of amyloid fibrils with a β-pleated sheet structure. Diagnosis typically relies on Congo red or Thioflavine T staining. Recently, DAPI (4',6-Diamidino-2-Phenylindole), which is a common nuclear fluorochrome, has been reported to stain amyloid. DAPI staining is simpler than Congo Red and Thioflavine T staining, but its staining mechanism remains unclear. Thus, this study aimed to investigate the mechanism and specificity of DAPI staining for amyloid and its utility. The staining properties of DAPI and Thioflavine T for amyloid were compared on the basis of their stereochemical similarity. In addition, formic acid-treated amyloid specimens were used to investigate the mechanism by which structural changes affect DAPI binding to amyloid fibrils. DAPI staining was also evaluated in four specimens with different types of amyloid. DAPI and Thioflavine T had similar stereochemistry and staining behavior. The amyloid present in formic acid-treated specimens was negative for DAPI staining, indicating that DAPI may recognize the conformation of amyloid fibrils. DAPI stained positively in specimens deposited with AA, Aβ, AL, and AIAPP. DAPI staining recognizes the β-pleated sheet structure of the amyloid fibril structure, and is a simple and sensitive method for detecting amyloid deposition.

Tartrazine, a coal tar-derived azo color, is utilized in food, drinks, cosmetics, and pharmaceuticals. Its azo group catabolizes in the gut, poisoning the liver. This study investigated the efficacy of melatonin, an endogenous antioxidant from the pineal gland against hepatotoxicity in tartrazine-intoxicated rats. Thirty-two adult male wistar albino rats were allocated into four groups: control group, melatonin group (10 mg/kg), tartrazine-treated group (7.5 mg/kg), and tartrazine + melatonin-treated group (7.5 mg/kg tartrazine + 10 mg/kg melatonin). Doses were taken daily for 4 weeks. Melatonin's influence on hepatotoxicity was assessed by monitoring liver enzyme activity, antioxidant state, apoptotic and inflammatory markers, DNA fragmentation, histological and ultrastructural changes. Rats exposed to tartrazine exhibited elevated liver enzymes, oxidant-antioxidant imbalance, and elevated hepatic inflammatory markers (TNF-α, IL-6). Tartrazine also damaged DNA and induced histological and ultrastructural alterations in liver tissue, as shown by the comet assay. Alpha-fetoprotein (AFP) and proliferating cell nuclear antigen (PCNA) were strongly expressed in immunohistochemistry. In rats, melatonin significantly reduced all tartrazine effects. Conversely, melatonin treatment significantly alleviated all aforementioned effects induced by tartrazine in rats by decreasing liver enzymes, elevating antioxidant enzymes, and reducing hepatic inflammatory markers. Enhanced histological assessment and the ultrastructure of the liver was detected following melatonin use. The use of melatonin may safeguard against tartrazine-induced hepatic DNA damage. In conclusion, the current findings indicate that tartrazine administration has detrimental health effects and deleterious impacts on liver function and structure. Melatonin mitigated tartrazine-induced liver damage via antioxidant, anti-inflammatory, and anti-apoptotic pathways.

Periodic acid-Schiff (PAS) staining is useful for visualizing glycogen and mucus. 7-amino-4-methylcoumarin (AMC), an organic reagent, exhibits blue fluorescent signals upon UV excitation. We recently showed that AMC can be used as a fluorescent substitute for Schiff's reagent in PAS staining. In-resin correlative light-electron microscopy (CLEM) is an excellent technique employing fluorescent and electron microscopy (EM) images obtained from the same resin-embedded ultra-thin sections, and provides a high level of morphological concordance between the fluorescent and EM images. Here we studied whether AMC was useful in detecting PAS-positive cells in in-resin CLEM using human pathological samples. Formalin-fixed, paraffin-embedded sections of human colonic and renal tissues and colonic amoebiasis were used. After staining with AMC, these samples were subjected to EM preparation and embedded in epoxy resin. Semi-thin and ultra-thin sections were prepared, and fluorescent and EM images were obtained. AMC signals were well preserved in colonic goblet cells, renal basement membrane, and amoebic bodies in epoxy resin-embedded sections. Using the CLEM technique, a small number of amoebic bodies were easily detected in an inflammatory background of colonic amoebiasis. In conclusion, we successfully detected PAS-positive cells in in-resin CLEM with AMC. Our method may enhance EM analysis in human pathology.

Safeguarding the integrity and functionality of the gastrointestinal system is paramount, given its vulnerability to several detrimental effects. One of the factors that can cause functional disorders is oxidative stress, which can disrupt the homeostasis of intestinal tissue and cause various diseases. In this study, we aimed to evaluate the expression patterns of nuclear factor erythroid 2-like 1 (Nrf1) and nuclear factor erythroid 2-like 2 (Nrf2) transcription factors, which are part of an important cleaning system that protects cells against oxidative stress, in the adult rat small intestine by immunohistochemical means. Six Wistar albino adult rats were used in this study. After taking 5 μm thick sections of the small intestine, immunohistochemistry labeling was performed to investigate the expression of Nrf1 and Nrf2. Both transcription factors were found to exhibit immunopositivity in the duodenum, jejunum, and ileum segments of the small intestine. In the crypt epithelium, Nrf1 and Nrf2 showed similar intensity and distribution of staining, whereas, in the villus epithelium, Nrf2 immunoreactivity was most prominent in the ileum and weakest in the jejunum. Nrf1 exhibited lower staining intensity in the ileum. In the smooth muscle layers and the myenteric plexus, Nrf2 immunopositivity was highest in the duodenum, while it was weaker in the jejunum and ileum. These findings indicate that Nrf1 and Nrf2 display region-specific expression patterns in the small intestine and suggest that these transcription factors may play distinct roles in the regional oxidative stress response of intestinal tissues.

The present study investigated the protective effect of Aloe vera extract against the toxic effects of doxorubicin (DOX) on mice ovaries cultured in vitro. Female Swiss mice with regular estrous cycle were used. The animals (n = 48) had their ovaries collected and cultured individually in a 24-well plate at 37.5°C in 5% CO₂ for 6 days. The ovaries were culture DMEM⁺ alone or supplemented with DOX (0.3 μg/ml), as well as both DOX and Aloe vera (5%, 10%, 25%, or 50%). After culture, ovaries were fixed for histological analysis (follicle morphology, growth and activation, extracellular matrix (ECM) configuration and stromal cell density), immunohistochemistry (TNF-α expression) or stored at - 80°C to evaluate the levels of mRNA for superoxide dismutase (SOD), catalase (CAT), nuclear factor-erythroid 2-related factor 2 (NRF2) and tumoral necrosis factor-α (TNF-α) by real-time PCR. The results showed that ovaries cultured with DOX had a lower percentage of normal follicles and reduced stromal cell density, but when Aloe vera extract was added to the culture medium, there was a protective effect on the ovarian structure against the deleterious effects of DOX. In addition, ovaries cultured with both DOX and Aloe vera (10 and 25%) had reduced TNF-α immunostaining and increased expression of mRNA for antioxidant enzymes (SOD, CAT, and NRF2). In conclusion, Aloe vera is associated with a protective effect on ovarian follicles and stromal cells against DOX-induced toxicity. Therefore, Aloe vera has great potential to preserve fertility in patients undergoing chemotherapy treatment, which often cause irreversible damage to oocytes.

Our study aimed to investigate the antidiabetic, antioxidative, and antiapoptotic effects of liraglutide on the testes of rats with diabetes and ischemia/reperfusion injury. Subjects were divided into three groups: control, diabetes, and torsion groups. Rats with diabetes were further divided into two subgroups such as diabetes and diabetes+Liraglutide groups. The torsion group was divided into three subgroups such as torsion, torsion/detorsion, and torsion/detorsion+Liraglutide groups. Malondialdehyde (MDA), superoxide dismutase (SOD), and testosterone levels were measured from blood samples. Also, testicular tissue samples were examined by light and electron microscopy. Apoptosis was assessed using immunohistochemistry for caspase-3. Degeneration of seminiferous tubules and interstitium was observed in the diabetes, torsion, and torsion/detorsion groups, while Liraglutide treated groups showed normal seminiferous tubules morphology. Elevated levels of apoptosis, i.e. caspase-3, were observed in diabetes, torsion, and torsion/detorsion groups (P < 0.05), whereas Liraglutide treated groups had similar levels of apoptosis as the control group. MDA levels of diabetes, torsion, and torsion/detorsion groups were increased (P < 0.05), while SOD and testosterone levels were decreased (P < 0.05). However, Liraglutide treated groups, SOD, MDA, and testosterone levels were found similar to the control group. In conclusion, Liraglutide positively affects structural changes and hormone levels in diabetes and torsion/detorsion groups.

Breakthrough progress has been made in the molecular mechanism research and clinical application of PD-1/PD-L1 in the regulation of immunosuppression and tolerance mainly in human and mouse fields, but is relatively slow in other species. The eukaryotic expression vectors pECFP-Fc-1 and pEYFP-Fc-L1 for high expression of canine PD-1 and PD-L1 proteins were constructed and transfected into human embryonic kidney 293 T cells. Fluorescence microscopy, laser scanning confocal microscopy, and immunofluorescence technology were used to identify the expression and membrane localization of the target proteins in human embryonic kidney 293 T cells. The binding activity of the target proteins expressed on the model cell was identified by eukaryotic expression vector co-transfection and immunocoprecipitation. The results showed that canine PD-1 and PD-L1 proteins were expressed on the membrane surfaces of their respective positively transfected cells. The cell membrane complex was further analyzed by co-immunoprecipitation technology, PD-L1 protein components were successfully detected in the pull-down complex of canine PD-1 antibody, and the two target proteins expressed in the model cells showed good mutual binding activity. Further research is needed to evaluate high throughput and a reliable method for screening drugs that block the PD-1 and PD-L1 pathway.

One of the most widely used techniques for cancer treatment is radiotherapy. Autophagic pathways allow some cancer cells that are resistant to radiation therapy to survive. Inhibiting autophagy has been shown to improve radiotherapy efficacy in several cancer types. Chloroquine (CQ) is a reasonable choice that has been used for many years to treat malaria and is preferred because of its minimal side-effects. Nevertheless, the effects of coadministration of CQ with radiation on various tissues remain unclear. In this study, it was aimed to understand how CQ, used to increase the effectiveness of radiotherapy, has effects on small intestine tissue alone and together with radiotherapy and what role apocynin (APO) can play with its antioxidant character in these stress conditions. Animals were divided into eight groups. The control group received physiological saline, while the other groups received 8 Gy total body irradiation, 50 mg/kg CQ, and 20 mg/kg APO, alone and in combination. In addition to causing significant histological damage, radiation triggered autophagy and showed antiproliferative and apoptotic effects. CQ administered with radiotherapy (RAD) had antiproliferative effects and did not cause a significant change in apoptosis. Reduced glutathione level, glutathione reductase, glutathione peroxidase, glutathione S-transferase, catalase, superoxide dismutase activities, and total antioxidant status were decreased, while lipid peroxidation, total oxidant status, reactive oxygen species, tumor necrosis factor-α, nitric oxide levels, adenosine deaminase, alkaline phosphatase, trypsin, lactate dehydrogenase, sodium potassium ATPase, xanthine oxidase activities, and protein carbonyl contents were increased in the RAD, CQ, and RAD+CQ groups. Apocynin therapy reversed these effects.

Liver ischemia and ischemia/reperfusion (I/R) injury are common in hepatic resection, trauma surgery, and transplantation and contribute to postoperative morbidity and mortality. This study aimed to investigate the relationship between early histopathological changes due to liver I/R injury and serum levels of perilipin-2 (Plin2) and other oxidative stress biomarkers. Fifty female Wistar albino rats were divided into five groups: Group 1 (control), Group 2 (ischemia for 30 minutes), and Groups 3-5 (ischemia followed by 1, 2, or 3 hours of reperfusion). Intracardiac and arterial blood samples were collected for biochemical analysis, while liver tissue samples were used for histopathological examination. Serum levels of Plin2, ischemia-modified albumin (IMA), total antioxidant status (TAS), and total oxidant status (TOS) were measured. Plin2 levels were significantly lower in the ischemia group compared to others (p < 0.01). The control group had significantly lower IMA, TOS, aspartate aminotransferase (AST), and alanine aminotransferase (ALT) levels (p < 0.01) and lower TAS levels (p < 0.05). Rats with Grade 1 liver injury showed significantly lower Plin2 (p < 0.01) and higher IMA levels (p < 0.05). Reduced serum Plin2 following ischemia suggests its potential as a biomarker for acute liver injury. Further studies are needed to clarify its role across different reperfusion durations.