Protective Effects of Mesenchymal Stem Cells Against Central Nervous System Injury in Heat Stroke.

BACKGROUND: Heatstroke (HS) is a serious disease caused by central nervous system (CNS) injuries, such as delirium, convulsion, and coma. Currently, mesenchymal stem cells (MSCs) have demonstrated novel neuroprotective effects; therefore, this research explores the neuroprotective effects and mechanisms of MSCs against HS injury. METHODS: HS rat models were induced in a 40°C and 65% humidity environment until the rectal temperature reached 42°C. The verified HS injury model rats were divided into the HS and MSCs-treated groups. Each rat in the treated group was infused with 1x106 MSCs suspended in 0.3 ml physiological saline via the tail vein. The HS- or MSCs-treated rats were further divided into early-stage (3d) and late-stage (28d). HS rat models were induced by a high-temperature and high-humidity environment at a specific time, the mortality was analyzed, and an automatic biochemical analyzer measured levels of liver and kidney function indicators in the blood. The neurons' morphologic changes were observed through Nissl staining, and neurological deficit scores were performed. Moreover, the levels of inflammatory factors in brain tissue were measured using a multi-cytokine detection platform, and the expression of BDNF, phosphorylated TrkB and P38 were detected by the Western Bolt. RESULTS: MSCs injection significantly reduced mortality and alleviated liver and kidney function. Moreover, the neurological deficit and neuronic edema of the hippocampus caused by HS at 3d and 28d were significantly ameliorated by MSCs administration. Specifically, the injection of MSCs inhibited high levels of interleukin (IL)-1ß, IL-6, tumor necrosis factor-α (TNF-α), and IL-17A caused by HS but elevated the levels of IL-10 and IL-13 in the early period (3d); while in the later period (28d), MSCs significantly increased the levels of IL-10 and IL-13 continuously and inhibited the high level of IL-17A. Furthermore, MSCs injection increased the expressions of BDNF and phosphorylated TrkB (BDNF receptor), meanwhile inhibiting the expression of phosphorylated P38 (inflammatory factor) in the brains of HS rats in the early period (3d) but had no significant influence on the later period (28d). CONCLUSION: These results suggested that MSCs injection may provide therapeutic effects for HS in rats by improving liver and kidney function and reducing CNS damage. Moreover, MSCs injection inhibited the brain inflammatory response of HS rats, and the BDNF-TrkB and P38/MAPK signal pathways may be involved, providing a potential mechanism for HS therapy by MSCs administration.

Role of ultrasonographic features and quantified BRAFV600E mutation in lymph node metastasis in Chinese patients with papillary thyroid carcinoma.

The association between cervical lymph node metastasis (LNM) and ultrasonographic features as well as BRAFV600E mutations in patients with papillary thyroid carcinoma (PTC) remained controversial. This study investigated the association between LNM and ultrasonographic features as well as BRAFV600E mutation in Chinese patients with PTC. A total of 280 patients with PTC in China were included in this study. 108 had cervical lymph node metastasis, while 172 had not. Younger age (<45years) and several ultrasonographic features were significantly associated with cervical LNM (Ps < 0.05). The BRAFV600E mutation was detected in 81.0% of patients with PTC (226/280). The status of BRAFV600E mutation was not associated with cervical LNM. However, Ct values by PCR and intensity of reactions by immunohistochemistry (IHC) for BRAFV600E expression had shown significant difference between group with and without LNM. Furthermore, an increased proportion of LNM was also found with the incremental intensity of IHC for BRAFV600E expression from weak to strong reaction after adjusted potential confounders. Further studies are required to verify this association and explore the intrinsic mechanism.

[The effect of rosuvastatin therapy on CCR2 expression in mononuclear cells and its upstream pathway].

OBJECTIVE: To investigate the effect of rosuvastatin therapy on C-C chemokine receptor(CCR2)expression in mononuclear cells in patients with carotid atherosclerosis and explore the possible upstream mechanism. METHODS: Twenty patients without previous statin treatment were enrolled. Rosuvastatin were given 5 to 20 mg/day for 3 months. At baseline and 12 weeks, lipid profile and plasma monocyte chemotactic protein-1 (MCP-1) levels were examined. The mRNA and protein expressions of CCR2 in the mononuclear cells were measured with RT-PCR and flow cytometry, respectively. The mRNA and protein expression of peroxidase proliferator-activated receptor(PPAR ß) were detected with RT-PCR and Western blot, respectively. RESULTS: After 3-months rosuvastatin treatment, the patients' low-density lipoprotein cholesterol (LDL-C) levels decreased significantly (P<0.01). Compared with baseline, the mRNA and protein expressions of CCR2 in the mononuclear cells showed significantly decrease, as well as plasma MCP-1 levels (P<0.05). Both mRNA and protein expressions of PPAR ß in the mononuclear cells increased (P<0.05). CONCLUSIONS: Rosuvastatin may attenuate MCP-1/CCR2 through PPARß upstream pathway.

[The effects of leptin on neuron apoptosis in mice with cerebral ischemia/reperfusion injury].

OBJECTIVE: To study the effect of leptin on neuron apoptosis in mice with cerebral ischemia injury. METHODS: Seventy-five male Kuming mice were randomly divided into 3 groups:sham, model and leptin intervention group, respectively. Focal cerebral ischemia/reperfusion injury model in mice was established by middle cerebral artery occlusion. Leptin intervention group was injected with leptin (1µg/g weight, I. P.) at 0 min of ischemic injury. Neuron apoptosis was detected by TUNEL staining. The mRNA expression of apoptosis relative gene bcl-2 and caspase-3 were detected by RT-PCR. The protein expression of bcl-2 and caspase-3 were detected by immunohistochemistry. RESULTS: In model group, most of the neurons in the central area of cerebral ischemia had necrosis obviously, and the amount of neuron apop-tosis was much higher than that in sham group (P<0.01). Compared with sham group, both expression of pro-apoptosis gene caspase-3 and anti-apoptosis gene bcl-2 increased significantly in model group (P<0.01). Compared with model group, the amount of neuron apoptosis and expression level of caspase-3 were decreased significantly (P<0.01), whereas the mRNA and protein expression of bcl-2 were increased sig-nificantly in leptin intervention group (P<0.01). CONCLUSIONS: Leptin could reduce neuron apoptosis through down-regulation the expression of caspase-3 and up-regulation the expression of bcl-2. It suggests that leptin could play a neuroprotective role in cerebral ischemia injury.

Inhibition of the connexin 43 elevation may be involved in the neuroprotective activity of leptin against brain ischemic injury.

Leptin is a multifunctional hormone produced by the ob gene and is secreted by adipocytes that regulate food intake and energy metabolism. Numerous studies demonstrated that leptin is a novel neuroprotective effector, however, the mechanisms are largely unknown. Herein, we demonstrate the protective activities of leptin after ischemic stroke and provide the first evidence for the involvement of the connexin 43 (Cx43) in leptin-mediated neuroprotection. We found that leptin treatment reduces the infarct volume, improves animal behavioral parameters, and inhibits the elevation of Cx43 expression in vivo. In vitro, leptin reverses ischemia-induced SY5Y and U87 cells Cx43 elevation, secreted glutamate levels in medium and SY5Y cell death, these roles could be abolished by leptin receptor blocker. Additionally, leptin administration upregulated the extracellular signal-regulated kinase1/2 (ERK1/2) phosphorylation. Moreover, ERK1/2 inhibitors pretreatment reversed the effects of leptin on Cx43 expression, glutamate levels and cell apoptosis. In conclusion, the present study demonstrated that leptin can reduce the Cx43 expression and cell death both in vivo and in vitro via ERK1/2 signaling pathway. This result provides a novel regulatory signaling pathway of the neuroprotective effects of leptin and may contribute to ischemic brain injury prevention and therapy.

Is leptin a predictive factor in patients with lung cancer?

OBJECTIVES: The aim of this study was to evaluate the expression and clinical significance of leptin in lung cancer. METHODS: 126 patients with lung cancer ranged from 30 to 83years of age were studied. Serum leptin levels were determined by ELISA. The mRNA and protein levels of leptin in normal and lung cancer tissues were measured by RT-PCR and immunohistochemistry. The relationships between leptin levels and clinicopathological factors were evaluated by Wilcoxon rank sum or Kruskal-Wallis H test. RESULTS: Serum leptin levels in lung cancer patients were significantly higher compared to those in controls and leptin expression in lung cancer tissue was markedly increased than that in normal lung tissue (both P<0.050). CONCLUSIONS: Determination of leptin levels might provide useful predictive information for lung cancer.

Localized leptin release may be an important mechanism of curcumin action after acute ischemic injuries.

BACKGROUND: Previous studies have demonstrated that both curcumin and leptin are protective factors against acute injuries. Here, we investigated whether leptin and its signaling pathway mediate the protective effects of curcumin. METHODS: A solid dispersion of curcumin-polyvinylpyrrolidone K30 was prepared and administered intraperitoneally. In vivo intestinal ischemia/reperfusion (I/R) injury in mice determined the effects of curcumin administration on inflammation, oxygen radical production, and leptin expression. In vitro studies using the venous epithelial cell line ECV-304 examined hypoxia/reoxygenation-induced leptin expression and release after curcumin administration. Furthermore, the effects on the leptin-regulated ERK1/2 and p38 MAPK signaling pathways were also explored. RESULTS: Intestinal I/R induced marked bowel injuries. Curcumin treatment significantly improved animal survival and reduced the pathologic injuries in the intestines. Furthermore, the elevated intestinal water content and levels of malondialdehyde, interleukin 1ß (IL-1ß) and IL-6 were significantly decreased, but levels of superoxide dismutase increased. Interestingly, we found that the decreased leptin and its receptor Ob-Rb were restored by curcumin administration. In addition, in vitro studies showed that curcumin increased leptin expression and release after hypoxia/reoxygenation-induced cell injuries. Moreover, curcumin treatment restored decreased ERK1/2 phosphorylation (p-ERK1/2) and inhibited overactive p38 (p-p38) after injuries, and the effect was reversed by a leptin-specific antibody or Ob-R blocker. CONCLUSION: These data suggest that leptin and Ob-Rb-dependent ERK and p38 MAPK signaling pathways may be involved in curcumin protection against intestinal I/R injury, and leptin may be a potential target of curcumin in intestinal I/R injury and other related acute diseases.

[The effect of leptin on Cx43 expression in protecting mice cerebral ischemia/reperfusion injury].

OBJECTIVE: To explore the effect of leptin on expression of Cx43 after rat cerebral ischemia/ reperfusion injury and its related mechanism. METHODS: Forty-five male kunming mice were randomly divided into 3 groups: sham group, model group and leptin group. Mouse models of transient focal cerebral ischemia were established by occlusion of the right middle cerebral artery for 2 h followed by 24 h reperfusion in model and leptin group. Mice of leptin group were intraperitoneally injected with 1 mg/kg leptin at 0 minute after ischemia. The infarct volume and neurological deficit scores following leptin treatment were determined using TTC staining and the Longa's score, respectively, to evaluate the protective effect of leptin against ischemic cerebral injury. The histopathological changes in the brain were observed with HE staining. The astrocytes of SD rat cerebral cotex were cultured primaryly and purified, and then divided them into four groups: control, model, leptin 100 microg/L, and leptin 500 microg/L. The cerebral astrocytes with hypoxia/reoxygenation injury were induced. The cellular viability of injury was detected by MTT assay. The effect of leptin on Cx43 expression was detected by Western blot in brain tissues and astrocytes. RESULTS: Compared with the model group, the neurological deficits and cerebral infarct volume of leptin group were reduced (P< 0.05), the histopathological injury in the brain tissues was alleviated and the expression of Cx43 was decreased markedly (P < 0.01). The survival rate of astrocytes was increased significantly in leptin 500 microg/L group (P < 0.01), whereas the Cx43 expression of astrocytes decreased (P < 0.01). But the difference of leptin 100 mcirog/L was not significant (P > 0.05). CONCLUSION: Leptin can ameliorate cerebral pathological changes in the event of IR injury by suppressing the expression of Cx43 both in vivo and vitro experiments.

Leptin administration alleviates ischemic brain injury in mice by reducing oxidative stress and subsequent neuronal apoptosis.

BACKGROUND: Recent research has indicates that leptin plays a protective role in traumatic brain injury. We studied the protective effect of leptin on cerebral ischemia/reperfusion injury by using mice transient focal cerebral ischemia/reperfusion injury model. METHODS: The distribution of 125I-leptin in the mouse brain was assessed by radioimmunoassay method. Mouse models of transient focal cerebral ischemia were established by occlusion of the right middle cerebral artery for two hours followed by 24 hours reperfusion. The neurologic deficits and infarct volume were determined using the Longa's score and 2,3,5-triphenyltetrazolium chloride staining, respectively. Regional cerebral blood flow was monitored by a laser-Doppler blood flowmeter. The levels of malondialdehyde, nitric oxide, nitric oxide synthase, and superoxide dismutase were detected according to respective assay kit. The histologic changes and neuronal apoptosis were observed with hematoxylin and eosin and transferase-mediated dUTP-biotin nick end labeling staining, respectively. The expression of B-cell lymphoma/leukemia-2 (Bcl-2) and cysteineasparateprotease-3 (caspase-3) were investigated by Western blot and real-time polymerase chain reaction assay. RESULTS: Leptin decreased infarct volume and neurologic defects and improved regional cerebral blood flow and microvascular branch blood flow after injury. The malondialdehyde and nitric oxide levels were reduced, and superoxide dismutase level was increased after leptin treatment, which also minimized histologic changes and neuronal apoptosis, led to the upregulation of Bcl-2 and downregulation of caspase-3 expression after injury. CONCLUSIONS: Peripherally administered leptin crossed the blood-brain barrier and was distributed into multiple regions of the brain; in the brain, leptin directly alleviated the injury-evoked damages by reducing oxidative stress and neuronal apoptosis.

Leptin relieves intestinal ischemia/reperfusion injury by promoting ERK1/2 phosphorylation and the NO signaling pathway.

BACKGROUND: Recently, research has indicated that leptin plays a protective role in traumatic brain and liver injury. We studied the protective effect of leptin on intestinal I/R injury and examined its mechanism by using mice intestinal I/R model and murine peritoneal macrophage hypoxia/reoxygenation (H/R) injury model. METHODS: Leptin was intraperitoneally administrated at 45 minutes after ischemia, then reperfusion for two hours. Cells were treated with different concentrations of leptin at three hours after hypoxia, then reoxygenation for six hours. Mice intestines were harvested for histopathologic properties. The malondialdehyde, nitric oxide (NO), interleukin-6, and total antioxidative capacity were detected according to respective assay kit. Phosphorylated extracellular regulated kinase1/2 (p-ERK1/2) and phosphorylated cytosolic phospholipase A(2) (p-cPLA2) were determined by Western blot assay. RESULTS: Here, we show that leptin reduced intestinal histologic alterations, malondialdehyde and interleukin-6 levels but increased the endogenous leptin expression and NO production in the intestines. Leptin also increased the NO and total antioxidative capacity levels in cells. We further demonstrated that leptin markedly activated ERK1/2 in the intestines and activated ERK1/2 and cPLA2 in the cells. Moreover, the protective effect of leptin against intestinal I/R injury and elevated NO production was attenuated by blocking the ERK1/2 pathway. CONCLUSIONS: These data demonstrate that leptin ameliorated intestinal I/R and peritoneal macrophage H/R injury by enhancing ERK1/2 phosphorylation and promoting the NO production signaling pathway.

Leptin attenuates cerebral ischemia/reperfusion injury partially by CGRP expression.

Ischemic stroke is a medical emergency triggered by a rapid reduction in blood supply to localized portions of the brain, usually because of thrombosis or embolism, which leads to neuronal dysfunction and death in the affected brain areas. Leptin is generally considered to be a strong and quick stress mediator after injuries. However, whether and how peripherally administered leptin performs neuroprotective potency in cerebral stroke has not been fully investigated. It has been reported that CGRP(8-37), an antagonist of the CGRP receptor, could reverse the protective effect of leptin on rats with CIP (caerulein-induced pancreatitis). However, the question remains: are leptin and CGRP associated in cerebral ischemia/reperfusion injury? The present study attempted to evaluate the relationship between CGRP expression and leptin neuroprotective effects (1mg/kg in 200 µL normal saline, i.p.) on focal cerebral ischemia/reperfusion injury in mice and the protective effect of leptin (500 µg/L) on neurons during hypoxia/reoxygenation injury. Peripheral administration of leptin alleviated injury-evoked brain damage by promoting CGRP expression, improving regional cerebral blood flow, and reducing local infarct volume and neurological deficits. Furthermore, leptin also promoted bcl-2 expression and suppressed caspase-3 in vivo and vitro after injury. Administration of CGRP(8-37) (4 × 10(-8)mol/L) partly abolished the beneficial effects of leptin, and restored the normal expression levels of bcl-2 and caspase-3 in neurons, which indicated that leptin-induced protection of neurons was correlated with release of CGRP. These results indicate that the neuroprotective effect of leptin against cerebral ischemia/reperfusion injury may be strongly relevant to the increase of CGRP expression.

[Correlation of killer immunoglobulin-like receptor gene diversity with nasopharyngeal carcinoma in Chinese southern Han population].

The objective of this study was to elucidate the correlation of killer immunoglobulin-like receptor (KIR) gene diversity with nasopharyngeal carcinoma (NPC) in the Chinese southern Han population. KIR genotyping of peripheral blood samples from 67 patients with NPC and 77 randomly-selected healthy controls was performed by PCR-SSP, the relative risk (RR) value was calculated by means of Wolf method. The results showed that the KIR2DL3 gene frequency in NPC patient group was significantly lower than that in healthy controls (χ²>3.84, p < 0.05, RR = 0.08), whereas the KIR2DS5 and KIR2DL5B gene frequencies in patient group were significantly higher than those in healthy controls (χ²>3.84, p < 0.05, RR > 1), the other KIR gene frequencies were no statistically different between two groups. It is concluded that the KIR2DL3, KIR2DS5 and KIR2DL5B genes may be correlated with pathogenesis of NPC in the Chinese southern Han population.

[The role of Leptin on neuron apoptosis in mice with cerebral ischemia/reperfusion injury].

OBJECTIVE: To study the effect of Leptin on neuron apoptosis in mice with cerebral ischemia injury and its mechanism. METHODS: Seventy-five mice were randomly divided into three groups. Focal cerebral ischemia/reperfusion injury model in mice was reproduced by middle cerebral artery occlusion for 2 hours followed by reperfusion. In Leptin intervention group mice were given Leptin 1 µg/g during cerebral ischemia by intraperitoneal injection. Mice in the model group were given equal amount of phosphate buffer saline. After reperfusion for 24 hours, the neuron apoptosis was detected by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining. The mRNA and protein expression of apoptosis relative gene caspase-3 and bcl-2 were determined by reverse transcription-polymerase chain reaction (RT-PCR) and immuno histochemistry. RESULTS: Most of neuron necrosis was observed in cerebral ischemia center in model group. Compared with sham-operation group, neuron apoptosis rate, mRNA and protein expression of caspase-3 and bcl-2 in model group increased significantly [apoptosis rate: (68.65 ± 0.79)% vs. (4.40 ± 0.00)%, caspase-3 mRNA: 2.563 ± 0.250 vs. 0.153 ± 0.020, bcl-2 mRNA: 0.337 ± 0.100 vs. 0.125 ± 0.030, caspase-3 protein (absorbance value, A value): 0.57 ± 0.05 vs. 0.37 ± 0.03, bcl-2 protein (A value): 0.51 ± 0.04 vs. 0.35 ± 0.01, all P<0.01]. The apoptosis rate of penumbra neurons was reduced in Leptin intervention group significantly compared with model group [(42.30 ± 8.45)% vs. (68.65 ± 0.79)%, P<0.01]. Compared with model group, the mRNA and protein expression of caspase-3 in Leptin intervention group were reduced significantly [caspase-3 mRNA: 2.267 ± 0.040 vs. 2.563 ± 0.250, caspase-3 protein (A value): 0.45 ± 0.04 vs. 0.57 ± 0.05, P>0.05 and P<0.01], and the mRNA and protein expression of bcl-2 in Leptin intervention group upregulated significantly [bcl-2 mRNA: 0.662 ± 0.040 vs. 0.337 ± 0.100, bcl-2 protein (A value): 0.76 ± 0.09 vs. 0.51 ± 0.04, both P<0.01]. CONCLUSION: Leptin could reduce apoptosis of neurons through down-regulation of the expression of caspase-3 and up-regulation of the expression of bcl-2. The results suggest that Leptin plays a neuroprotective role in cerebral ischemia injury.

[Effects of leptin on apoptosis of rat cerebral astrocytes with ischemia/hypoxia injury].

OBJECTIVE: To investigate the effect of leptin on apoptosis of rat cerebral astrocytes with ischemia/ hypoxia injury and its mechanism. METHODS: The cerebral astrocytes with ischemic/hypoxia injury were induced in neonatal SD rats. The cellular viability of injury of astrocytes was detected by MTT assay. The apoptosis of astrocyte were detected with Annexin V-FITC kit. The effect of leptin on the expression of apoptosis factor bcl-2, bax, caspase-3 was detected by RT-PCR and Western blot. RESULTS: Compared with the ischemia group, the cellular viability of leptin intervention group increased significantly (P < 0.01), while the astrocytes apoptosis of leptin intervention group decreased significantly (P < 0.01). The mRNA and protein expression level of antiapoptosis factor bcl-2 in leptin intervention group was much higher than that of ischemia group (P < 0.01), whereas the mRNA and protein expression of bax and caspase-3 was much lower than that of ischemia group (P < 0.01). CONCLUSION: Leptin could significantly decrease the apoptosis of astrocytes with ischemia/hypoxia injury, and it i relevant to the increase of bcl-2 expression and the decrease of bax caspase-3 expression level.

[Effects of ethyl pyruvate on injuries of sepsis in mice].

OBJECTIVE: To explore the effect of ethyl pyruvate (EP) and alkaline phosphatase (ALP) on injuries of sepsis and the mechanism involved. METHODS: A murine sepsis model of cecal ligation and puncture was reproduced, and 90 male Kunming mice were divided randomly into sham-operation, model and EP-intervention groups. 75 mg/kg EP was intraperitoneally injected in EP groups 1 hour after establishment of model, and the mice in model group were given a same volume of Ringer's solution. The eyeballs were removed in the latter two groups, and mice were sacrificed at 15 minutes and 1, 3 and 6 hours in subgroups of 10 mice each. ALP, uric acid (UA) and ratio of lactic acid and pyruvic acid were determined in serum and homogenized lung tissue by autonomous biochemical analyzer, and pathological changes in intestine were observed by hematoxylin-eosin (HE) staining. RESULTS: Compared with sham-operation group, serum ALP in model groups and EP groups decreased significantly (P<0.05 or P<0.01), and ALP level of EP group was significantly lower than model group at 6 hours after injury (P<0.05). Compared with sham-operation group, serum UA in model group increased significantly at 1 hour, and reached the highest level at 3 hours (both P<0.05) but decreased significantly later. UA in EP group was significantly lower than that in model group at 1 hour and 3 hours (both P<0.05). Lactic acid/pyruvic acid ratio in lung homogenate of EP group was significantly lower than that of the model group at all the time points (all P<0.05). Intestinal structural damages were distinctly improved in EP group compared with model group at 3 hours and 6 hours (both P<0.05 ). CONCLUSION: EP promotes the utilization of serum ALP, decreases serum UA, ameliorates acidosis and intestinal damages, thus exerting a protective effect on sepsis-induced organ injuries.