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Situ Cell Apoptosis Detection Kit I


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  • Dose-dependent immunohistochemical changes in rat cornea and retina after oral methylphenidate administration. 19007354

    Methylphenidate hydrochloride (MPH), more commonly known as Ritalin, is a piperidine derivative and is the drug most often used to treat attention deficit/hyperactivity disorder, one of the most common behavioural disorders of children and young adults. The aim of this study was to investigate dose-dependent immunohistochemical Dopamine 2 receptor (D2) expression and apoptosis in the rat cornea and cornea. In this study, 27 female pre-pubertal Wistar albino rats, divided into three different dose groups (5, 10 and 20 mg/kg) and their control groups, were used. They were treated orally with methylphenidate dissolved in saline solution for 5 days per week during 3 months. At the end of the third month, after perfusion fixation, eye tissue was removed. Paraffin sections were collected for immunohistochemical and terminal deoxynucleotidyl-transferase-mediated dUTP-biotin nick end labelling assay studies. In our study, we observed that the cornea D2 receptor reactivity showed a dose-related increase after MPH treatment, especially in basal cells of the epithelium and a dose-dependent decrease in the retinal ganglion cell which was statistically meaningful. Analysis of the cornea thickness results showed no meaningful difference between groups. Apoptotic cell number showed a meaningful increase in the high dose treated group compared to the other groups of the study. The data suggest that Ritalin has degenerative effect on the important functional part of the eye, such as cornea and retina and its activating dopaminergic mechanism via similar neuronal paths, functionally and structurally, to induce morphological changes. As a result, we believe that this morphological changes negatively effecting functional organization of the affected cornea and retina.
    Document Type:
    Reference
    Product Catalog Number:
    S7101
    Product Catalog Name:
    ApopTag® Plus Peroxidase In Situ Apoptosis Kit

  • Effect of fluoride intoxication on endometrial apoptosis and lipid peroxidation in rats: role of vitamins E and C. 17227693

    Fluoride is a strong, hard anion and cumulative toxic agent. The effect of fluoride intoxication on lipid peroxidation in endometrial tissue and the protective effects of combinations of vitamins E and C in rats were studied. Additionally, the apoptotic changes in endometrial tissue were examined. Experimental groups were as follows: control group; a group treated with 100 mg/l fluoride (F group); and a group treated with 100 mg/l fluoride plus vitamins E and C (F+Vit group). The F and F+Vit groups were treated orally with fluoride for 30 days. Vitamins E and C were injected simultaneously at doses of 50 mg/kg day i.m. and 20 mg/kg day body weight i.p. Extensive formation of DNA strand breaks, the typical biochemical feature of apoptosis, was detected with the use of the terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-biotin nick and labeling (TUNEL) method. Malondialdehyde (MDA) levels were determined in uterine tissue of rats. Fluoride caused a significant increase in MDA levels (an important marker of lipid peroxidation) in the fluoride group compared with the controls (p0.05). Vitamins E and C significantly reduced the fluoride-induced lipid peroxidation in the F+Vit group compared with the F group (p0.05). Diffuse apoptosis in glandular epithelium and stromal cells was found in endometrial tissues of F treated rats by TUNEL method. The severity of these lesions was reduced by the administration of vitamins. From these results, it can be concluded that subchronic fluoride administration causes endometrial apoptosis, and lipid peroxidation may be a molecular mechanism involved in fluoride-induced toxicity. Furthermore, treatment with a combination of vitamins E and C reduced endometrial apoptosis caused by fluoride.
    Document Type:
    Reference
    Product Catalog Number:
    S7101
    Product Catalog Name:
    ApopTag® Plus Peroxidase In Situ Apoptosis Kit

  • Serine/threonine phosphatase (SP-STP), secreted from Streptococcus pyogenes, is a pro-apoptotic protein. 22262847

    This investigation illustrates an important property of eukaryote-type serine/threonine phosphatase (SP-STP) of group A Streptococcus (GAS) in causing programmed cell death of human pharyngeal cells. The secretory nature of SP-STP, its elevated expression in the intracellular GAS, and the ability of wild-type GAS but not the GAS mutant devoid of SP-STP to cause apoptosis of the host cell both in vitro and in vivo suggest that GAS deploys SP-STP as an important virulence determinant to exploit host cell machinery for its own advantage during infection. The exogenously added SP-STP is able to enter the cytoplasm and subsequently traverses into the nucleus in a temporal fashion to cause apoptosis of the pharyngeal cells. The programmed cell death induced by SP-STP, which requires active transcription and de novo protein synthesis, is also caspase-dependent. Furthermore, the entry of SP-STP into the cytoplasm is dependent on its secondary structure as the catalytically inactive SP-STP with an altered structure is unable to internalize and cause apoptosis. The ectopically expressed wild-type SP-STP was found to be in the nucleus and conferred apoptosis of Detroit 562 pharyngeal cells. However, the catalytically inactive SP-STP was unable to cause apoptosis even when intracellularly expressed. The ability of SP-STP to activate pro-apoptotic signaling cascades both in the cytoplasm and in the nucleus resulted in mitochondrial dysfunctioning and perturbation in the phosphorylation status of histones in the nucleus. SP-STP thus not only functions as a virulence regulator but also as an important factor responsible for host-related pathogenesis.
    Document Type:
    Reference
    Product Catalog Number:
    S7100
    Product Catalog Name:
    ApopTag® Peroxidase In Situ Apoptosis Detection Kit

  • Absence of glutathione peroxidase 4 affects tumor angiogenesis through increased 12/15-lipoxygenase activity. 20234819

    The selenoenzyme glutathione peroxidase 4 (GPx4) has been described to control specific cyclooxygenases (COXs) and lipoxygenases (LOXs) that exert substantiated functions in tumor growth and angiogenesis. Therefore, we hypothesized a putative regulatory role of GPx4 during tumor progression and created transformed murine embryonic fibroblasts with inducible disruption of GPx4. GPx4 inactivation caused rapid cell death in vitro, which could be prevented either by lipophilic antioxidants or by 12/15-LOX-specific inhibitors, but not by inhibitors targeting other LOX isoforms or COX. Surprisingly, transformed GPx4(+/-) cells did not die when grown in Matrigel but gave rise to tumor spheroids. Subcutaneous implantation of tumor cells into mice resulted in knockout tumors that were indistinguishable in volume and mass in comparison to wild-type tumors. However, further analysis revealed a strong vascular phenotype. We observed an increase in microvessel density as well as a reduction in the number of large diameter vessels covered by smooth muscle cells. This phenotype could be linked to increased 12/15-LOX activity that was accompanied by an up-regulation of basic fibroblast growth factor and down-regulation of vascular endothelial growth factor A protein expression. Indeed, pharmacological inhibition of 12/15-LOX successfully reversed the tumor phenotype and led to normalized vessel morphology. Thus, we conclude that GPx4, through controlling 12/15-LOX activity, is an important regulator of tumor angiogenesis as well as vessel maturation.
    Document Type:
    Reference
    Product Catalog Number:
    S7100
    Product Catalog Name:
    ApopTag® Peroxidase In Situ Apoptosis Detection Kit

  • Heat shock protein 70 (HSP70) is critical for the photoreceptor stress response after retinal detachment via modulating anti-apoptotic Akt kinase. 21356360

    Photoreceptor apoptosis is a major cause of vision loss in many ocular diseases. Significant progress has been made to elucidate the molecular pathways involved in this process, yet little is known about proteins counteracting these apoptotic pathways. It is established that heat shock proteins (HSPs) function as molecular helper proteins (chaperones) by preventing protein aggregation and facilitating refolding of dysfunctional proteins, critical to the survival of all organisms. Here, we investigated the role of HSP70 on photoreceptor survival after experimental retinal detachment (RD) in mice and rats. We found that HSP70 was up-regulated after RD and associated with phosphorylated Akt, thereby preventing its dephosphorylation and further activation of cell death pathways. Administration of quercetin, which inhibits HSP70 and suppresses Akt phosphorylation significantly increased photoreceptor apoptosis. Similarly, RD-induced photoreceptor apoptosis was augmented in mice carrying hypomorphic mutations of the genes encoding HSP70. On the other hand, administration of geranylgeranylacetone, which induces an increase in HSP70 significantly decreased photoreceptor apoptosis after RD through prolonged activation of Akt pathway. Thus, HSP70 may be a favorable potential target to increase photoreceptor cell survival after RD.
    Document Type:
    Reference
    Product Catalog Number:
    S7110
    Product Catalog Name:
    ApopTag® Fluorescein In Situ Apoptosis Detection Kit

  • Jmjd1a demethylase-regulated histone modification is essential for cAMP-response element modulator-regulated gene expression and spermatogenesis. 19910458

    Spermatogenesis, a fundamental process in the male reproductive system, requires a series of tightly controlled epigenetic and genetic events in germ cells ranging from spermatogonia to spermatozoa. Jmjd1a is a key epigenetic regulator expressed in the testis. It specifically demethylates mono- and di-methylated histone H3 lysine 9 (H3K9me1 and H3K9me2) but not tri-methylated H3K9 (H3K9me3). In this study, we generated a Jmjd1a antibody for immunohistochemistry and found Jmjd1a was specifically produced in pachytene and secondary spermatocytes. Disruption of the Jmjd1a gene in mice significantly increased H3K9me1 and H3K9me2 levels in pachytene spermatocytes and early elongating spermatids without affecting H3K9me3 levels. Concurrently, the levels of histone acetylation were decreased in Jmjd1a knock-out germ cells. This suggests Jmjd1a promotes transcriptional activation by lowering histone methylation and increasing histone acetylation. Interestingly, the altered histone modifications in Jmjd1a-deficient germ cells caused diminished cAMP-response element modulator (Crem) recruitment to chromatin and decreased expression of the Crem coactivator Act and their target genes Tnp1 (transition protein 1), Tnp2, Prm1 (protamine 1), and Prm2, all of which are essential for chromatin condensation in spermatids. In agreement with these findings, Jmjd1a deficiency caused extensive germ cell apoptosis and blocked spermatid elongation, resulting in severe oligozoospermia, small testes, and infertility in male mice. These results indicate that the Jmjd1a-controlled epigenetic histone modifications are crucial for Crem-regulated gene expression and spermatogenesis.
    Document Type:
    Reference
    Product Catalog Number:
    Multiple
    Product Catalog Name:
    Multiple

  • Open blunt crush injury of different severity determines nature and extent of local tissue regeneration and repair. 20069568

    Insufficiency of skeletal muscle regeneration is often accompanied with functional deficiencies. The goal of our study was to assess the restoration of peripheral muscle upon injury of different severity. Blunt crush injury of the soleus muscle in rats was induced by a clamp and stepwise amplified in severity by rising the locking level of the clamp, resulting in three different groups (1x lock; 2x lock; 3x lock; n = 30 animals per group). After assessment of the fast twitch and tetanic contraction capacity at days 1, 4, 7, 14, and 42 postinjury sampling of muscle tissue served for analysis of cell proliferation, including satellite cells, apoptosis, and leukocyte infiltration. Contraction force analysis demonstrated significantly higher values of relative muscle strength in the 1x lock group compared to the two other groups over 42 days. Calculation of the twitch-to-tetanic force ratio revealed significantly higher mean values at days 1, 7, and 14 in the animals of group 2x lock and 3x lock, indicating a transformation toward a fast-twitching muscular phenotype. Moreover, cell proliferation during the first 4 days was found dependent on the severity of muscle injury in that the higher the severity the higher the proliferation. At the same time, cell apoptosis was found increased, and at day 1 the local leukocyte infiltration was significantly higher in the 3x lock compared to the 1x lock group. These data indicate that severity of injury correlates with local repair responses, which, however, are not necessarily sufficient to fully restore muscle function. (c) 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.
    Document Type:
    Reference
    Product Catalog Number:
    S7100
    Product Catalog Name:
    ApopTag® Peroxidase In Situ Apoptosis Detection Kit

  • Fibroblast growth factor-2-overexpressing myoblasts encapsulated in alginate spheres increase proliferation, reduce apoptosis, induce adipogenesis, and enhance regenerati ... 21815803

    The fibroblast growth factor 2 (FGF-2) is known as pleiotropic cytokine with myoblast proliferative properties. In the present study, we tested the hypothesis that gene transfer of human FGF-2 via transplantation of genetically modified L8-myoblast encapsulated in alginate modulates the skeletal muscle recovery after crush injury in Wistar rats. Therefore, we performed a crush injury to the soleus muscle and transplanted alginate spheres containing myoblasts genetically modified to overexpress human FGF-2 (FGF-2) or a luciferase (LUC) cDNA at the site of injury. Animals that underwent muscle injury without transplantation of alginate spheres served as control (control). At day 4 after trauma the FGF-2 group showed significant higher mean values of cell proliferation (bromodeoxyuridine immunohistochemistry) and significant lower values of cell apoptosis (terminal deoxynucleotidyl transferase nick end labeling histology) compared to animals receiving luciferase-overexpressing myoblasts. At the same time point adiponectin expression (ACRP30 immunohistochemistry) was increased in the FGF-2 group exclusively. The p75(NTR) expression (p75(NTR) immunohistochemistry) significantly improved in both the FGF-2 and LUC group compared to the control group. Functional analysis of the injured muscle did not reveal a significant increase of the muscle force in the FGF-2 group compared to the control and LUC group 14 days after injury. In vitro analysis for 14 days of the FGF-2-modified spheres demonstrated at day 7 and day 14 a significant increase of the relative cell count as well as of the relative viable cell count in the FGF-2 myoblast spheres compared to luciferase myoblast spheres. Additionally, the expression of FGF-2 (enzyme-linked immunosorbent assay analysis) and luciferase (chemiluminescence analysis) persisted in vitro for 4 and 14 days, respectively. These results demonstrate that FGF-2-overexpressing myoblasts cannot considerably improve muscle strength but are able to modulate the proliferation as well as the apoptosis of injured muscle tissue mainly by conducting adipogenesis.
    Document Type:
    Reference
    Product Catalog Number:
    S7100
    Product Catalog Name:
    ApopTag® Peroxidase In Situ Apoptosis Detection Kit

  • Effects of erythropoietin in skin wound healing are dose related. 19403513

    The hematopoietic growth factor erythropoietin (EPO) attracts attention due to its all-tissue-protective pleiotropic properties. We studied the effect of EPO on dermal regeneration using intravital microscopy in a model of full dermal thickness wounds in the skin-fold chamber of hairless mice. Animals received repetitive low doses or high doses of EPO (RLD-EPO or RHD-EPO) or a single high dose of EPO (SHD-EPO). SHD-EPO accelerated wound epithelialization, reduced wound cellularity, and induced maturation of newly formed microvascular networks. In contrast, RHD-EPO impaired the healing process, as indicated by delayed epithelialization, high wound cellularity, and lack of maturation of microvascular networks. Also, RHD-EPO caused an excessive erythrocyte mass and rheological malfunction, further deteriorating vessel and tissue maturation. Moreover, RHD-EPO altered fibroblast and keratinocyte migration in vitro, while both cell types exposed to RLD-EPO, and, in particular, to SHD-EPO showed accelerated wound scratch closure. In summary, our data show that a single application of a high dose of EPO accelerates and improves skin wound healing.
    Document Type:
    Reference
    Product Catalog Number:
    S7101
    Product Catalog Name:
    ApopTag® Plus Peroxidase In Situ Apoptosis Kit

  • Effects of resveratrol on blood homocysteine level, on homocysteine induced oxidative stress, apoptosis and cognitive dysfunctions in rats. 22995369

    We aimed to examine the protective effects of resveratrol against homocysteine induced oxidative stress, apoptosis and cognitive impairment. Rats were randomly divided into three groups. Control group received standard rat food; homocysteine group (Hcy group) received daily methionine at a dose of 1g/kg-body weight dissolved in drinking water for thirty days; third group (Hcy+Res group) received same amount of methionine plus 20mg/kg/day resveratrol intraperitoneally for thirty days. Cognitive performances of the animals were tested by Morris water maze test. Then all animals were sacrificed to study lipid peroxidation (LPO), DNA fragmentation and p53 mRNA expression in the rat brain. The aortas of the sacrificed rats were processed for histopathological examination. Apoptosis in the aortas was assessed by TUNEL staining. Resveratrol significantly decreased serum levels of homocysteine, reversed Hcy induced LPO increase, decreased DNA fragmentation and p53 mRNA expression in the rat brains, and improved homocysteine induced impairment of long term spatial memory. Resveratrol could inhibit homocysteine induced apoptosis and histopathological deterioration in the rat aortic sections. In conclusion, resveratrol is effective in preventing homocysteine induced vascular and neural defects. In hyperhomocysteinemic rat model, our findings consequently warrant in future studies to reveal the true improvement mechanism of resveratrol.
    Document Type:
    Reference
    Product Catalog Number:
    S7101
    Product Catalog Name:
    ApopTag® Plus Peroxidase In Situ Apoptosis Kit