Blockade of L-Type Ca2+ Channel Activity Alleviates Oligodendrocyte Pathology following Brain Injury in Male Rats.

A growing body of studies suggests that Ca2+ signaling controls a variety of biological processes in brain elements. Activation of L-type voltage-operated Ca2+ channels (VOCCs) plays a role in the development of oligodendrocyte (OL) lineage loss, and indicates that the blocking of these channels may be an effective way to inhibit OL lineage cell loss. For this study, 10.5-day-old male Sprague-Dawley rats were used to generate cerebellar tissue slices. The slice tissues were cultured and randomly allocated to one of four groups (six each) and treated as follows: Group I, (sham control); Group II, 0.1% dimethyl sulfoxide (DMSO) only (vehicle control); Group III, injury (INJ); Group IV, (INJ and treatment with NIF). The injury was simulated by exposing the slice tissues to 20 min of oxygen-glucose deprivation (OGD). At 3 days post-treatment, the survival, apoptosis, and proliferation of the OL lineages were measured and compared. Results: In the INJ group, there was a decrease in mature myelin basic protein+ OLs (MBP+ OLs) and their precursors, NG2+ OPCs (Nerve-glia antigen 2+ oligodendrocyte precursor cell), compared with controls. A significant elevation was observed in the NG2+ OPCs and apoptotic MBP+ OLs as confirmed by a TUNEL assay. However, the cell proliferation rate was decreased in NG2+ OPCs. NIF increased OL survival as measured by apoptosis rate in both OL lineages and preserved the rate of proliferation in the NG2+ OPCs. Conclusions: Activation of L-type VOCCs may contribute to OL pathology in association with reduced mitosis of OPCs following brain injury as a strategy to treat demyelinating diseases.

Therapeutic potential of vitamin D against bisphenol A-induced spleen injury in Swiss albino mice.

Bisphenol A (BPA), a ubiquitous plasticizer, is capable of producing oxidative splenic injury, and ultimately led to spleen pathology. Further, a link between VitD levels and oxidative stress was reported. Hence the role of VitD in BPA-induced oxidative splenic injury was investigated in this study. Sixty male and female Swiss albino mice (3.5 weeks old) were randomly divided into control and treated groups 12 mice in each (six males and six females). The control groups were further divided into sham (no treatment) and vehicle (sterile corn oil), whereas the treatment group was divided into VitD (2,195 IU/kg), BPA (50 µg/kg), and BPA+VitD (50 µg/kg + 2,195 IU/kg) groups. For six weeks, the animals were dosed intraperitoneally (i.p). One week later, at 10.5 weeks old, mice were sacrificed for biochemical and histological analyses. Findings showed BPA triggered neurobehavioral abnormalities and spleen injury with increased apoptotic indices (e.g. DNA fragmentation) in both sexes. A significant increase was found in lipid peroxidation marker, MDA in splenic tissue, and leukocytosis. Conversely, VitD treatment altered this scenario into motor performance preservation, reducing oxidative splenic injury with a decrease in the percent apoptotic index. This protection was significantly correlated with preserving leukocyte counts and reduced MDA levels in both genders. It can be concluded from the above findings that VitD treatment has an ameliorative effect on oxidative splenic injury induced by BPA, highlighting the continuous crosstalk between oxidative stress and the VitD signaling pathway.

Vitamin D ameliorates liver pathology in mice caused by exposure to endocrine disruptor bisphenol A.

Background: Increasing evidence suggests that bisphenol A (BPA) induces liver pathological changes. Further, an association between BPA and circulating vitamin D (VitD) levels were documented. Aim: The role of VitD in BPA-induced liver pathological changes was explored in this study. Methods: Healthy 4.5-week-old male (n = 35) and female (n = 35) Swiss albino mice were used in this study. The animals were randomly divided into control and treated groups. The control groups were further divided into sham (no treatment) and vehicle (corn oil), whereas the treated groups were also divided into VitD (2195 U/kg), BPA (50 µg/kg), and BPA + VitD (50 µg/kg + 2195 U/kg) groups. For 6 weeks (twice a week), the animals were dosed intraperitoneally. One week later (at 10.5-weeks-old), the animals were sacrificed for biochemical and histological analyses. Results: BPA produced a considerable rise in the body and liver weights in both genders of mice when compared to control mice. BPA also caused significant increases in the liver damage markers alanine transaminase (ALT), alkaline phosphatase (ALP), and gamma-glutamyl transferase (GGT). It also induced liver histopathological changes, including higher apoptotic indices in both genders. On the other hand, treatment with VitD considerably reduced liver damage and slightly decreased the apoptotic index rate. The ALP, ALT, and GGT levels were also markedly reduced. VitD has been proven to have a protective effect on both genders. Conclusions: According to our findings, VitD protects mice from BPA-induced liver damage, possibly via suppressing liver damage markers.

Nigella sativa Oil Alleviates Mouse Testis and Sperm Abnormalities Induced by BPA Potentially through Redox Homeostasis.

BACKGROUND & AIM: Significant evidence indicates that endocrine disrupted bisphenol A (BPA) seriously endangers human health. In males, BPA affects testis architecture and sperm quality, and ultimately reduces fertility. This study explored the therapeutic potential of Nigella sativa (NS) seed extract on testis and sperm abnormalities in BPA-exposed mice and characterized the underlying mechanism. METHODS: Forty male Swiss albino mice (5.5 weeks old, N = 8 per group) were randomly divided into five groups: Group I, normal control, Group II, vehicle control (sterile corn oil); Group III, NS-exposed (oral 200 mg/kg); Group IV, BPA-exposed (oral 400 µg/kg body weight); Group V, BPA + NS-exposed mice. Animals were treated for 6 weeks and sacrificed for biochemical and histological examination. RESULTS: The results indicated that BPA exposure results in significant testis and sperm abnormalities. Specifically, BPA promoted a marked reduction in the body and testis compared with the control group. Histopathological findings showed that BPA caused a widespread degeneration of spermatogenic cells of the seminiferous epithelium, decreased sperm counts and motility, and augmented sperm abnormalities, and whereas little alteration to sperm DNA was observed. In addition, BPA increased the levels of the lipid peroxidation marker, malondialdehyde (MDA), and reduced the levels of the antioxidant marker, reducing glutathione (GSH). Treatment with NS oil extract during BPA exposure significantly alleviated testis and sperm abnormalities, reduced MDA levels, and enhanced GSH levels. CONCLUSION: The results demonstrate that NS oil protects mice against BPA-induced sperm and testis abnormalities, likely by suppressing levels of the oxidative stress marker, MDA, and enhancing the levels of the antioxidant marker, GSH.

Histone deacetylase 2 inhibitor valproic acid attenuates bisphenol A-induced liver pathology in male mice.

Accumulating evidence indicates the role of endocrine disruptor bisphenol A (BPA) in many pathological conditions. Histone deacetylase (HDAC) inhibition has potential for the treatment of many diseases/abnormalities. Using a mouse BPA exposure model, this study investigated the hepatoprotective effects of the Food and Drug Administration-approved HDAC2 inhibitor valproic acid (VPA) against BPA-induced liver pathology. We randomly divided 30 adult male Swiss albino mice (8 weeks old; N = 6) into five groups: group 1, no treatment (sham control (SC)); group 2, only oral sterile corn oil (vehicle control (VC)); group 3, 4 mg/kg/day of oral BPA (single dose (BPA group)); group 4, 0.4% oral VPA (VPA group); and group 5, oral BPA + VPA (BPA + VPA group). At the age of 10 weeks, the mice were euthanized for biochemical and histological examinations. BPA promoted a significant decrease in the body weight (BW), an increase in the liver weight, and a significant increase in the levels of liver damage markers aspartate aminotransferase and alanine aminotransferase in the BPA group compared to SC, as well as pathological changes in liver tissue. We also found an increase in the rate of apoptosis among hepatocytes. In addition, BPA significantly increased the levels of oxidative stress indices, malondialdehyde, and protein carbonylation but decreased the levels of reduced glutathione (GSH) in the BPA group compared to SC. In contrast, treatment with the HDAC2 inhibitor VPA significantly attenuated liver pathology, oxidative stress, and apoptosis and also enhanced GSH levels in VPA group and BPA + VPA group. The HDAC2 inhibitor VPA protects mice against BPA-induced liver pathology, likely by inhibiting oxidative stress and enhancing the levels of antioxidant-reduced GSH.

Blocking of NF­kB/p38 MAPK pathways mitigates oligodendrocyte pathology in a model of neonatal white matter injury.

Reactive gliosis and inflammation are risk factors for white matter injury (WMI) development, which are correlated with the development of many neurodevelopmental deficits with no treatment. This study aimed to understand the mechanisms correlated with WMI, with a particular focus on the role of nuclear factor­kappa B (NF­kB) and p38 mitogen­activated protein kinases (MAPKs) pathways. Seven­day­old Wistar rats were used to generate cerebellar tissue slices. Slices were cultured and randomly allocated to one of 3 groups and treated as follows: group­I (control); group­II (WMI), slices were subjected to 20 min of oxygen­glucose deprivation (OGD); group­III (WMI+ blockers), slices were subjected to OGD and treated with the blockers. Results showed that OGD insult triggered a marked increase in the apoptosis among WM elements, as confirmed by TUNEL assay. Immunocytochemical experiments revealed that there was a significant decrease in the percent of MBP+ OLs and NG2+ OPCs, and myelin integrity. There was also a significant increase in the percent of reactive microglia and astrocytes. BrdU immunostaining revealed there was an increase in the percent of proliferating microglia and astrocytes. Q­RT­PCR results showed OGD upregulated the expression levels of cytokines (TNF­α, IL­1, IL­6, and IL­1ß) and inducible nitric oxide synthase (iNOS). On the other hand, treatment with BAY11 or SB203580 significantly enhanced the OL survival, restored myelin loss, and reduced microglia and astrocyte reactivity, and downregulated the iNOS and cytokine expression. Our findings demonstrate that blocking of NF­KB/p38 MAPK pathways alleviated reactive gliosis, inflammation, and OL loss upon WMI. The findings may help to develop therapeutic interventions for WMI.

Exposure to low-dose bisphenol A induces spleen damage in a murine model: Potentially through oxidative stress?

Background: During early life, exposure to environmental toxicants, including endocrine disruptor bisphenol A (BPA), can be detrimental to the immune system. To our knowledge, a few researches have looked at the effects of developing BPA exposures on the spleen. Aim: The murine model was developed to investigate the underlying molecular mechanisms and mode of BPA actions on the spleen subsequent to prolonged early-life exposure to BPA. Methods: Immature (3-week-old) male and female Swiss Albino mice were intraperitoneally injected with 50 µg/kg BPA in corn oil or corn oil alone for 6 weeks. Mouse spleens were harvested and examined histologically at 10 weeks old (adulthood). Results: We observed neurobehavioral impairments and a significant increase in peripheral monocyte and lymphocyte counts in mice (males and females). Moreover, several spleen abnormalities in both male and female mice were observed in adulthood. BPA-treated mice's histopathological results revealed toxicity in the form of significantly active germinal centers of the white pulp and a few apoptotic cells. There was also a notable invasion of the red pulp by eosinophils and lymphocytes that were significantly higher than normal. Agarose gel electrophoresis provided further evidence of internucleosomal DNA fragmentation and apoptosis in the splenic tissues of BPA-treated mice compared to controls. In addition, there were increased levels of the lipid peroxidation malondialdehyde end-product, a marker of oxidative lipid damage, in the spleens of BPA-treated mice compared to controls. Conclusion: Our study provides evidence that oxidative stress injury induced by early-life exposures to BPA could contribute to a range of splenic tissue damages during adulthood.

Vitamin D mitigates adult onset diseases in male and female mice induced by early-life exposure to endocrine disruptor BPA.

Background: During early development, environmental compounds can induce adult onset diseases and disrupt the circulating vitamin D (VitD) levels. Aim: This study aimed to examine the protective role of VitD against the adverse effects of BPA on male and female mice. Methods: A total of 60 male and female Swiss Albino mice (3 weeks old) were randomly divided into 5 groups; each consisted of 12 mice (6 males and 6 females) and was treated as follows: Group I received no treatment (sham control); Group II, sterile corn oil only (vehicle control); Group III, BPA (400 µg/kg); Group IV, VitD (2,195 IU/kg); and Group V, BPA + VitD. At 10.5 weeks, the animals were sacrificed to conduct histological examinations. Results: BPA-exposed mice were found to have neurobehavioral abnormalities, heart, kidney, and lung diseases with increased apoptotic indices in both sexes. On the other hand, the treatment of BPA mice with VitD altered this scenario with modulated motor activity, enhanced body and organ weights, and preserved the heart, kidney, and lung architecture, alongside a decreased percent apoptotic index. Conclusion: Our findings illustrate that VitD protects mice against BPA-induced heart, kidney, and lung abnormalities.

HDAC2/3 inhibitor MI192 mitigates oligodendrocyte loss and reduces microglial activation upon injury: A potential role of epigenetics.

Background: During development, oligodendrocyte (OL) lineage cells are susceptible to injury, leading to life-long clinical neurodevelopmental deficits, which lack effective treatments. Drugs targeting epigenetic modifications that inhibit histone deacetylases (HDACs) protect from many clinical neurodegenerative disorders. Aim: This study aimed to investigate the therapeutic potential of histone deacetylase 2/3 (HDAC2/3) inhibitor MI192 on white matter (WM) pathology in a model of neonatal rat brain injury. Methods: Wistar rats (8.5-day-old, n = 32) were used to generate brain tissues. The tissues were cultured and then randomly divided into four groups and treated as following: group I (sham); the tissues were cultured under normoxia, group II (vehicle); DMSO only, group III (injury, INJ); the tissues were exposed to 20 minutes oxygen-glucose deprivation (OGD) insult, and group IV (INJ + MI192); the tissues were subjected to the OGD insult and then treated with the MI192 inhibitor. On culture day 10, the tissues were fixed for biochemical and histological examinations. Results: The results showed that inhibition of HDAC2/3 activity alleviated WM pathology. Specifically, MI192 treatment significantly reduced cell death, minimized apoptosis, and mitigates the loss of the MBP+ OLs and their precursors (NG2+ OPCs). Additionally, MI192 decreased the density of reactive microglia (OX-42+). These findings demonstrate that the inhibition of HDAC2/3 activity post-insult alleviates WM pathology through mechanism(s) including preserving OL lineage cells and suppressing microglial activation. Conclusion: The findings of this study suggest that HDAC2/3 inhibition is a rational strategy to preserve WM or reverse its pathology upon newborn brain injury.

Inhibition of ionotropic GluR signaling preserves oligodendrocyte lineage and myelination in an ex vivo rat model of white matter ischemic injury.

Preterm infants have a high risk of neonatal white matter injury (WMI). WMI leads to reduced myelination, inflammation, and clinical neurodevelopmental deficits for which there are no effective treatments. Ionotropic glutamate receptor (iGluR) induced excitotoxicity contributes to oligodendrocyte (OL) lineage cell loss and demyelination in brain models of neonatal and adult WMI. Here, we hypothesized that simulated ischemia (oxygen­glucose deprivation) damages white matter via activation of iGluR signaling, and that iGluR inhibition shortly after WMI could mitigate OL loss, enhance myelination, and suppress inflammation in an ex vivo cerebellar slice model of developing WMI. Inhibition of iGluR signaling by a combined block of AMPA and NMDA receptors, shortly after simulated ischemia, restored myelination, reduced apoptotic OLs, and enhanced OL precursor cell proliferation and maturation as well as upregulated expression of transcription factors regulating OL development and remyelination. Our findings demonstrate that iGluR inhibition post­injury alleviates OL lineage cell loss and inflammation and promotes myelination upon developing WMI. The findings may help to develop therapeutic interventions for the WMI treatment.

Selective adenosine A2A receptor inhibitor SCH58261 reduces oligodendrocyte loss upon brain injury in young rats.

Cellular elements of maturing brain are vulnerable to insults, which lead to neurodevelopmental defects. There are no established treatments at present. Here we examined the efficacy of selective adenosine A2A receptor inhibitor SCH58261 to combat brain injury, particularly oligodendrocyte (OL) lineage cells, in young rats. Wistar rats (n = 24, 6.5 days old) were randomly divided into equal groups of four. The sham (SHAM) group received no treatment, the vehicle (VEHICLE) group received 0.1% dimethylsufoxide, the injury (INJ) group was exposed to oxygen-glucose deprivation insult, and the injury+SCH58261 (INJ+SCH58261) group was exposed to the insult and received 1 µM SCH58261. Immunocytochemical experiments revealed that there was a significant reduction in the populations of mature OL (MBP+ OLs) and immature OL precursors (NG2+ OPCs) in the INJ group compared to SHAM group. Furthermore, there was also a significant increase in the percent of apoptotic MBP+ OL and NG2+ OPC populations as evidenced by TUNEL assay. In addition, there was a significant reduction in the proliferation rate among NG2+ OPCs, which was confirmed by BrdU immunostaining. On the other hand, treatment with SCH58261 significantly enhanced survival, evidenced by the reduction in apoptotic indices for both cell types, and it is preserved the NG2+ OPC proliferation. Activation of adenosine A2A receptors may contribute to OL lineage cell loss in association with decreased mitotic behavior of OPCs in neonatal brains upon injury. Future investigations assessing ability of SCH58261 to regenerate myelin will provide insights into its wider clinical relevance.

Paternal bisphenol A exposure induces testis and sperm pathologies in mice offspring: Possibly due to oxidative stress?

Bisphenol A (BPA), an endocrine and metabolic disruptor, is widely used to manufacture polycarbonate plastics and epoxy resins. Accumulating evidence suggests that paternal BPA exposure adversely affects male germlines and results in atypical reproductive phenotypes that might persist for generations to come. Our study investigated this exposure on testicular architecture and sperm quality in mouse offspring, and characterised underlying molecular mechanism(s). A total of 18 immature male Swiss albino mice (3.5 weeks old) were randomly divided into three groups and treated as follows: Group I, no treatment (sham control); Group II, sterile corn oil only (vehicle control); Group III, BPA (400 µg/kg) in sterile corn oil. At 9.5 weeks old, F0 males were mated with unexposed females. F0 offspring (F1 generation) were monitored for postnatal development for 10 weeks. At 11.5 weeks old, the animals were sacrificed to examine testicular architecture, sperm parameters, including DNA integrity, and oxidative stress biomarkers. Results showed that BPA significantly induced changes in the body and testis weights of the F0 and F1 generation BPA lineages compared to F0 and F1 generation control lineages. A decrease in sperm count and motility with further, increased sperm abnormalities, no or few sperm DNA alterations and elevated levels of MDA, PC and NO were recorded. Similar effects were found in BPA exposed F0 males, but were more pronounced in the F0 offspring. In addition, BPA caused alterations in the testicular architecture. These pathological changes extended transgenerationally to F1 generation males' mice, but the pathological changes were more pronounced in the F1 generation. Our findings demonstrate that the biological and health BPA impacts do not end in paternal adults, but are passed on to offspring generations. Hence, linking observed testis and sperm abnormalities in the F1 generation to BPA exposure of their parental line was evident in this work. The findings also illustrate that oxidative stress appears to be a molecular component of the testis and sperm pathologies.

Effects of storage temperature on the quantity and integrity of genomic DNA extracted from mice tissues: A comparison of recovery methods.

Efficient extraction of genomic DNA (gDNA) from biological materials found in harsh environments is the first step for successful forensic DNA profiling. This study aimed to evaluate two methods for DNA recovery from animal tissues (livers, muscles), focusing on the best storage temperature for DNA yield in term of quality, quantity, and integrity for use in several downstream molecular techniques. Six male Swiss albino mice were sacrificed, liver and muscle tissues (n=32) were then harvested and stored for one week in different temperatures, -20°C, 4°C, 25°C and 40°C. The conditioned animal tissues were used for DNA extraction by Chelex-100 method or NucleoSpinC Blood and Tissue kit. The extracted gDNA was visualized on 1.5% agarose gel electrophoresis to determine the quality of gDNA and analysed spectrophotometrically to determine the DNA concentration and the purity. Both methods, Chelex-100 and NucleoSpin Blood and Tissue kit found to be appropriate for yielding high quantity of gDNA, with the Chelex 100 method yielding a greater quantity (P < 0.045) than the kit. At -20°C, 4°C, and 25°C temperatures, the concentration of DNA yield was numerically lower than at 40°C. The NucleoSpinC Blood and Tissue kit produced a higher (P=0.031) purity product than the Chelex-100 method, particularly for muscle tissues. The Chelex-100 method is cheap, fast, effective, and is a crucial tool for yielding DNA from animal tissues (livers, muscles) exposed to harsh environment with little limitations.

Environmentally toxicant exposures induced intragenerational transmission of liver abnormalities in mice.

Environmental toxicants such as chemicals, heavy metals, and pesticides have been shown to promote transgenerational inheritance of abnormal phenotypes and/or diseases to multiple subsequent generations following parental and/or ancestral exposures. This study was designed to examine the potential transgenerational action of the environmental toxicant trichloroethane (TCE) on transmission of liver abnormality, and to elucidate the molecular etiology of hepatocyte cell damage. A total of thirty two healthy immature female albino mice were randomly divided into three equal groups as follows: a sham group, which did not receive any treatment; a vehicle group, which received corn oil alone, and TCE treated group (3 weeks, 100 µg/kg i.p., every 4th day). The F0 and F1 generation control and TCE populations were sacrificed at the age of four months, and various abnormalities histpathologically investigated. Cell death and oxidative stress indices were also measured. The present study provides experimental evidence for the inheritance of environmentally induced liver abnormalities in mice. The results of this study show that exposure to the TCE promoted adult onset liver abnormalities in F0 female mice as well as unexposed F1 generation offspring. It is the first study to report a transgenerational liver abnormalities in the F1 generation mice through maternal line prior to gestation. This finding was based on careful evaluation of liver histopathological abnormalities, apoptosis of hepatocytes, and measurements of oxidative stress biomarkers (lipid peroxidation, protein carbonylation, and nitric oxide) in control and TCE populations. There was an increase in liver histopathological abnormalities, cell death, and oxidative lipid damage in F0 and F1 hepatic tissues of TCE treated group. In conclusion, this study showed that the biological and health impacts of environmental toxicant TCE do not end in maternal adults, but are passed on to offspring generations. Hence, linking observed liver abnormality in the offspring to environmental exposure of their parental line. This study also illustrated that oxidative stress and apoptosis appear to be a molecular component of the hepatocyte cell injury.