MicroRNA-135a participates in the development of astrocytes derived from bacterial meningitis by downregulating HIF-1α.

Accumulating evidence has highlighted the potential of microRNAs (miRs) as biomarkers in various human diseases. However, the roles of miRs in bacterial meningitis (BM), a severe infectious condition, still remain unclear. Thus, the present study aimed to investigate the effects of miR-135a on proliferation and apoptosis of astrocytes in BM. Neonatal rats were injected with Streptococcus pneumoniae to establish the BM model. The expression of miR-135a and hypoxia-inducible factor 1α (HIF-1α) in the BM rat models were characterized, followed by determination of their interaction. Using gain- and loss-of-function approaches, the effects of miR-135a on proliferation, apoptosis, and expression of glial fibrillary acidic protein (GFAP), in addition to apoptosis-related factors in astrocytes were examined accordingly. The regulatory effect of HIF-1α was also determined along with the overexpression or knockdown of HIF-1α. The results obtained indicated that miR-135a was poorly expressed, whereas HIF-1α was highly expressed in the BM rat models. In addition, restored expression levels of miR-135a were determined to promote proliferation while inhibiting the apoptosis of astrocytes, along with downregulated Bax and Bad, as well as upregulated Bcl-2, Bcl-XL, and GFAP. As a target gene of miR-135a, HIF-1α expression was determined to be diminished by miR-135a. The upregulation of HIF-1α reversed the miR-135a-induced proliferation of astrocytes. Taken together, the key findings of the current study present evidence suggesting that miR-135a can downregulate HIF-1α and play a contributory role in the development of astrocytes derived from BM, providing a novel theoretical perspective for BM treatment approaches.

Clinical study of cerebral palsy in 408 children with periventricular leukomalacia.

This study aimed to investigate the high risk factors, cerebral palsy (CP) subtypes and comorbidities of periventricular leukomalacia (PVL). Based on treatment conditions at a specialist hospital, a cross-sectional clinical study and retrospective analysis of computed tomography and magnetic resonance imaging examinations was conducted to evaluate the risk factors, subtypes and comorbidities of CP in children with PVL. Among the 408 children with PVL, 8.58% were born with a weight of ≤1,500 g and 44.36% were born with a weight of ≥2,500 g. In addition, 36.76% of these children had a gestational age of ≤32 weeks and 37.75% had a gestational age of ≥37 weeks. The proportion of the children born with various high risk factors was 95.59%, including perinatal infections and hypoxia. Severe PVL was observed in preterm infants (63.41% with a gestational age of <28 weeks and 21.95% with a gestational age of 28-30 weeks) and low-birth weight infants, which were prone to quadriplegia (43.90%). The common comorbidities included visual and auditory disorders, epilepsy, mental retardation and language barriers. Visual and auditory disorders (26.96%) were the most common comorbidities. PVL was identified primarily in premature and low-birth weight infants. The degree of PVL was found to be negatively correlated with gestational age and birth weight. The degree of PVL in the full-term infants correlated with exposure to infections or hypoxia. Quadriplegia is common among the various subtypes of CP. Visual and hearing disorders are the most common comorbidities of CP; these comorbidities occurred most frequently with quadriplegia.

Upregulation of the TPX2 gene is associated with enhanced tumor malignance of esophageal squamous cell carcinoma.

PURPOSE: To explore the expression of TPX2 and its significance in esophageal squamous cell carcinoma (ESCC) tissue and approach relationship between the TPX2 and clinicopathological characteristic of esophageal squamous cell carcinoma. METHOD: RT-PCR and immunohistochemical staining were used to compare the expression of TPX2 in 62 esophageal squamous cell carcinoma, 31 atypical hyperplasia and 62 normal esophageal mucosa. RESULTS: In ESCC, atypical hyperplasia and in normal mucous membrane tissues, the positive rate of TPX2 protein expression was 85.5% (53/62), 51.6% (16/31) and 4.8% (3/62); the positive rate of TPX2 mRNA expression was 65.5% (40/62), 35.5 (11/31) and 4.83% (3/62). The expression of TPX2 protein and mRNA were correlated with invasive depth and lymphatic metastasis of ESCC (P<0.01). CONCLUSIONS: Overexpression of TPX2 may be risk factor of lymph node in esophageal carcinoma, and maybe a potential biomarker for early diagnosis and prognosis of esophageal squamous cell carcinoma.

Sulfated beta-glucan derived from oat bran with potent anti-HIV activity.

China is a major producer of oats; the annual harvested area of 350,000 ha yields approximately 465,000 tons, giving an average yield of 1.33 tons/ha. The bran is not used for animal feed as it is of poor digestibility and low nutritive content and is considered a waste byproduct. Therefore, it is advantageous to produce a value-added product from the bran. We extracted the native polysaccharide, a linear (1-3)-, (1-4)-linked beta-glucan (OBG) from the oat bran and synthesized a sulfated derivative OBGS containing 36.5% sulfate. OBGS had potent activity against a primary isolate of human immunodeficiency virus (HIV-1) in peripheral blood mononuclear cells at a concentration (EC(50)=5.98 x 10(-4) microM) approximately 15,000 times below its cytotoxic concentration. OBGS was also active postinfection (EC(50)=5.3 x 10(-4) microM) and protected pretreated peripheral mononuclear cells (EC(50)=5.2 x 10(-2) microM) washed free of the compounds prior to infection. Thus, OBGS has potential as a vaginal microbicide and is the first such report for oat bran derived sulfated beta-glucan.

Changes of cell proliferation and differentiation in the developing brain of mouse.

OBJECTIVE: To investigate the cell proliferation and differentiation in the developing brain of mouse. METHODS: C57/BL6 mice were divided into 3 groups at random. Bromodeoxyuridine (BrdU) was injected into the brains in different development periods once a day for 7 d. The brains were retrieved 4 weeks after the last BrdU injection. Immunohistochemical and immunofluorescent studies were carried out for detecting cell proliferation (BrdU) and cell differentiation (NeuN, APC, Iba1, and S100beta), respectively. RESULTS: The number of BrdU labeled cells decreased significantly with the development of the brain. Cell proliferation was prominent in the cortex and striatum. A small portion of BrdU and NeuN double labeled cells could be detected in the cortex at the early stage of development, and in the striatum and CA of the hippocampus in all groups. The majority of BrdU labeled cells were neuroglia, and the number of neuroglia cells decreased dramatically with brain maturation. Neurogenesis is the major cytogenesis in the dentate gyrus. CONCLUSION: These results demonstrated that cell proliferation, differentiation and survival were age and brain region related.

[Neuroprotective effect of hypothermia on hypoxic-ischemic brain injury in mice].

OBJECTIVE: The study was to investigate the effect of different temperatures during hypoxia on brain injury in mice of different ages. METHODS: Newborn C57/BL6 mice at 7 days or 21 days of life were subjected to left carotid artery ligation followed by exposure with 10% oxygen. The mice were kept in a incubator with a predetermined, constant temperature, either 34 degrees centigrade (Hypothermia group) or 36 degrees centigrade (Normothermia group). Brain injury was evaluated 7 days after hypoxia-ischemia (HI). Active caspase-3 and apoptosis-inducing factor (AIF) expressions in the brain tissue were detected by immunohistochemistry and Western Blot was used to evaluate the phosphor-Akt (P-Akt) expression in the brain tissue at 24 hrs post-HI. RESULTS: Brain injuries, including the cortex, hippocampus, striatum and thalamus injuries, occurred in the Normothermia group at 7 days post-HI. The brain cortex showed cystic cavitation in the postnatal day (P)7 pups mice and laminar infarct of the brain cortex was observed in P21 mice. In the Hypothermia group, the P7 mice did not present with laminar infarct of the cortex and had lower scores of neuropathological lesions in cortex, hippocampus, striatum and thalamus than P7 mice from the Normothermia group (P < 0.01); the cortex injuries were significantly relieved but the injuries of hippocampus, striatum and thalamus in P21 mice were similar to those from the Normothermia group. Active caspase-3 (7.0 +/- 5.6) and AIF positive cells (3.7 +/- 6.2) in the cortex of P7 mice from the Hypothermia group were significantly lower than those of the Normothermia group (51.5 +/- 23.2 and 31.8 +/- 22.4) at 24 hrs post-HI (P < 0.01). Wetstern Blot showed the P-Akt expression was obviously decreased in the ipsilateral hemisphere to the occlusion compared with that of the contralateral hemisphere after HI in the Normothermia group (P < 0.05), while in the Hypothermia group the P-Akt expression was not significantly different between the two hemispheres. CONCLUSIONS: Hypothermia has protective effects against HI insults. The protection was more pronounced for the immature brain than the mature brain.

[Expression of p53 in neonatal mice following hypoxia-ischemia and effects of its inhibitor on neonatal brain injury].

OBJECTIVE: p53-induced apoptosis is crucial in the development of hypoxic-ischemia (HI) brain damage and neurodegenerative disorders. Some experimental research has shown that a synthetic inhibitor of p53 can protect neurons against apoptosis. This study aimed to explore the expression of p53 in neonatal mice following HI brain damage and the effect of p53 inhibitor (pifithrin-alpha, PFT-alpha) on brain damage. METHODS: HI was induced in 9-day-old mice pups by ligation of left carotid artery and 10% oxygen exposure for 55 minutes. The pups were sacrificed and the brains were taken out at 3, 8, 24, and 72 hrs post-HI. The brains were sectioned and stained with antibody against p53 and microtubule-associated protein 2 (MAP-2). PFT-alpha was injected intraperitoneally: in experiment 1, immediately after HI with different dosages (1, 2 and 8 mg/kg); in experiment 2, 2 mg/kg at different HI times (1 hr before HI, and immediately and 1 hr after HI). Control animals without HI received injections of 0.5% dimethyl sulfoxide. Brain damage was evaluated by gross morphology scoring at 72 hrs after HI. RESULTS: The number of p53 positive cells in the cortex, hippocampus and striatum of the ipsilateral hemisphere increased significantly and peaked at 3-8 hrs post-HI when compared with those of contralateral hemisphere as well as normal controls. The positive cells distributed mainly in the MAP-2 negative area. Both different dosages and different injection time PFT-alpha treatment did not reduce the extent of brain damage. CONCLUSIONS: The immunoactivity of p53 increased significantly as early as 3 hrs post-HI. The distribution area of p53 expression was consistent with that of brain damage. The p53 inhibitor PFT-alpha has no protective effects against HI brain damage in neonatal mice.

Developmental changes of glutamate acid decarboxylase-67 in mouse brain after hypoxia ischemia.

Objective To study the developmental changes of glutamic acid decarboxylase-67 (GAD-67, a GABA synthetic enzyme) in normal and hypoxic ischemic (HI) brain. Methods C57/BL6 mice on postnatal day (P) 5, 9, 21and 60, corresponding developmentally to premature, term, juvenile and adult human brain were investigated by using both Western blot and immunohistochemistry methods either in normal condition or after hypoxic ischemic insult. Results The immunoreactivity of GAD-67 was up regulated with brain development and significant difference was seen between mature (P21, P60) and immature (P5, P9) brain. GAD-67 immunoreactivity decreased in the ipsilateral hemisphere in all the ages after hypoxia ischemia (HI) insult, but, significant decrease was only seen in the immature brain. Double labeling of GAD-67 and cell death marker, TUNEL, in the cortex at 8h post-HI in the P9 mice showed that (15.6 +/- 7.0)%TUNEL positive cells were GAD-67 positive which was higher than that of P60 mice. Conclusion These data suggest that GABAergic neurons in immature brain were more vulnerable to HI insult than that of mature brain.

[Cytochrome C release and apoptosis in neonatal rat cerebral hypoxia-ischemia].

OBJECTIVE: To study the relation of cytochrome C release from mitochondria to cytosol and neuronal apoptosis after cerebral hypoxia-ischemia (HI) in neonatal rats. METHODS: Hypoxia-ischemia was induced in 7-day-old rat pups by ligation of left carotid artery and 7.7% oxygen was inhaled for 55 min. The pups were sacrificed and the brains were taken out at different recovery time. Some of the brains were homogenized and cellular fraction of mitochondria and cytosol was isolated with different speed centrifugation. The cellular fraction was used for Western blotting. Some of the brains were sectioned and stained with antibody against cytochrome C and TUNEL as well as double labeling with different combinations. RESULTS: Western blots showed that cytochrome C in mitochondria was not reduced significantly at 1 h, but reduced markedly at 14 h in ipsilateral hemisphere post-HI. However, the immunoreactivity of cytochrome C in cytosol was increased markedly at 1 h post-HI and reached peak at 14 h post-HI. The number of cytochrome C positive cells in the cortex was increased significantly at 1 h (8.4 +/- 1.8/visual field) compared to normal control (1.5 +/- 0.8/visual field) (P < 0.01) and reached peak at 14 h (29.0 +/- 5.2/visual field) post-HI. The number of TUNEL positive cells increased significantly at 1 h post-HI (14 +/- 3/visual field) compared to normal control (1.5 +/- 0.8/visual field) (P < 0.01) and reached peak at 24 h (286 +/- 86/visual field). The double labeling of cytochrome C and active caspase-3 showed that they colocalized well at 3 h after HI. Furthermore, the positive cells showed nuclei condensation. There were more active caspase-3 positive cells at late recovery (24 h and on) after HI. The double labeling of cytochrome C and TUNEL showed only part of Positive cells colocalized. The cells with cytochrome C strong staining showed TUNEL negative or weakly positive. The cells with TUNEL strong staining showed weakly cytochrome C staining. CONCLUSION: Cytochrome C release is one of the early biochemical changes of neuronal apoptosis after hypoxia-ischemia in neonatal rat brain.

[Neuroprotective effect and mechanisms of hypothermia in neonatal rat cerebral hypoxic-ischemic damages].

OBJECTIVE: Recent studies suggest that hypothermia may be a potential treatment for perinatal hypoxic-ischemic (HI) brain damage. But the mechanisms of this effect are not well known. In the present study, the protective effect of systemic hypothermia as well as effect on apoptosis and associated biochemical events were investigated on neonatal rats with HI brain damage. METHODS: Seven-day-old Wistar rats were subjected to left carotid artery ligation and hypoxia was persisted for 60 min. Immediately at the end of hypoxia, the animals were maintained either at 36 degrees C or 30 degrees C for 10 h at random. Caspase-2, 3 activity in brain homogenate was detected with Western blotting at 24 h post-HI (n = 8 for each group). Immunoactivity of microtubule-associated protein-2 (MAP-2), active caspase-3, apoptosis inducing factor (AIF) and oligonucleotide hairpin probe staining were detected at 72 h post-HI. The infarct volume, neuronal loss in CA(1) sector of hippocampus as well as brain injury scoring were calculated according to MAP-2 staining and hematoxylin and eosin staining. RESULTS: Caspase-2, 3 activities were much higher in the normothermia group [(27.7 +/- 14.7), (94.9 +/- 53.1) pmol/(min.mg protein)] at 24 h post-HI than those of hypothermia [(7.9 +/- 3.4), (21.1 +/- 18.7) pmol/(min.mg protein)] and normal control groups [(7.6 +/- 0.7), (12.9 +/- 0.5) pmol/(min x mg protein)] (P < 0.01). The activities were not significantly different between hypothermia group and normal control group. Western blotting showed that caspase-3 activation process was blocked by hypothermia. The number of active caspase-3 and AIF positive cells in the cortex of ipsilateral hemisphere was much higher in the normothermia group (median: 148.5; 22/field) than that of hypothermia group (median: 48.5; 9/field) (P < 0.05). The number of apoptotic cells as judged by oligonucleotide hairpin probe labeling was much higher in normothermia group (median: 144/field) than that of hypothermia group (median: 133/field) (P < 0.05). The brain injury scoring, infarct volume and neuronal loss in CA(1) area of hippocampus were much less in the hypothermia group [10.4 +/- 2.9; 40.5 +/- 34.8)mm(3); 25.7 +/- 11.5] than that of normothermia group [14.2 +/- 3.5; (73.9 +/- 22.4) mm(3); 37.4 +/- 10.6, P < 0.05]. CONCLUSIONS: Systemic hypothermia for 10 h after hypoxia-ischemia seemed to be effective in reducing brain damage and the mechanism is associated with alteration of apoptotic pathway.