Treadmill exercise exerts a synergistic effect with bone marrow mesenchymal stem cell-derived exosomes on neuronal apoptosis and synaptic-axonal remodeling.

Treadmill exercise and mesenchymal stem cell transplantation are both practical and effective methods for the treatment of cerebral ischemia. However, whether there is a synergistic effect between the two remains unclear. In this study, we established rat models of ischemia/reperfusion injury by occlusion of the middle cerebral artery for 2 hours and reperfusion for 24 hours. Rat models were perfused with bone marrow mesenchymal stem cell-derived exosomes (MSC-exos) via the tail vein and underwent 14 successive days of treadmill exercise. Neurological assessment, histopathology, and immunohistochemistry results revealed decreased neuronal apoptosis and cerebral infarct volume, evident synaptic formation and axonal regeneration, and remarkably recovered neurological function in rats subjected to treadmill exercise and MSC-exos treatment. These effects were superior to those in rats subjected to treadmill exercise or MSC-exos treatment alone. Mechanistically, further investigation revealed that the activation of JNK1/c-Jun signaling pathways regulated neuronal apoptosis and synaptic-axonal remodeling. These findings suggest that treadmill exercise may exhibit a synergistic effect with MSC-exos treatment, which may be related to activation of the JNK1/c-Jun signaling pathway. This study provides novel theoretical evidence for the clinical application of treadmill exercise combined with MSC-exos treatment for ischemic cerebrovascular disease.

Six-month follow-up of functional status in discharged patients with coronavirus disease 2019.

BACKGROUND: The long-term functional outcome of discharged patients with coronavirus disease 2019 (COVID-19) remains unresolved. We aimed to describe a 6-month follow-up of functional status of COVID-19 survivors. METHODS: We reviewed the data of COVID-19 patients who had been consecutively admitted to the Tumor Center of Union Hospital (Wuhan, China) between 15 February and 14 March 2020. We quantified a 6-month functional outcome reflecting symptoms and disability in COVID-19 survivors using a post-COVID-19 functional status scale ranging from 0 to 4 (PCFS). We examined the risk factors for the incomplete functional status defined as a PCFS > 0 at a 6-month follow-up after discharge. RESULTS: We included a total of 95 COVID-19 survivors with a median age of 62 (IQR 53-69) who had a complete functional status (PCFS grade 0) at baseline in this retrospective observational study. At 6-month follow-up, 67 (70.5%) patients had a complete functional outcome (grade 0), 9 (9.5%) had a negligible limited function (grade 1), 12 (12.6%) had a mild limited function (grade 2), 7 (7.4%) had moderate limited function (grade 3). Univariable logistic regression analysis showed a significant association between the onset symptoms of muscle or joint pain and an increased risk of incomplete function (unadjusted OR 4.06, 95% CI 1.33-12.37). This association remained after adjustment for age and admission delay (adjusted OR 3.39, 95% CI 1.06-10.81, p = 0.039). CONCLUSIONS: A small proportion of discharged COVID-19 patients may have an incomplete functional outcome at a 6-month follow-up; intervention strategies are required.

Treadmill exercise enhances therapeutic potency of transplanted bone mesenchymal stem cells in cerebral ischemic rats via anti-apoptotic effects.

BACKGROUND: The transplantation of bone marrow stromal cells (MSCs) has proved to ameliorate ischemic brain injury in animals, but most transplanted MSCs undergo apoptosis in the ischemic penumbra, greatly compromising the therapeutic value of this treatment. Meanwhile, cell apoptosis can be inhibited by post-ischemia exercise which has been demonstrated to improve the expression of related anti-apoptotic proteins. The present study investigated whether treadmill exercise enhances the neuroprotective effects of transplanted MSCs in a rat experimental stroke model. RESULT: Rats were subjected to 2-h middle cerebral artery occlusion (MCAO). Twenty-four hours after reperfusion, they were assigned randomly to receive no MSCs treatment and no exercise (control group), intravenous transplantation of MSCs and treadmill exercise (MSCs + Ex group), MSCs transplantation only (MSCs group) and treadmill exercise only (Ex group). Neurological assessment, TUNEL staining and western blot were performed. Compared with the MSCs group and Ex group, the MSCs + Ex group reported markedly improved neurological function, significantly decreased apoptotic cells, and increased expressions of survivin and bcl-2 (p < 0.05 or p < 0.01, respectively). Interestingly, the treadmill exercise significantly inhibited the apoptosis of transplanted MSCs. As a result, the number of engrafted MSCs in the MSCs + Ex group was significantly higher than that in the MSC group (p < 0.01). CONCLUSIONS: Treadmill exercise enhances the therapeutic potency of MSCs by improving neurological function and possibly inhibiting the apoptosis of neuron cells and transplanted MSCs. These effects may involve an increased expression of survivin and bcl-2.

Correlating interleukin-10 promoter gene polymorphisms with human cerebral infarction onset.

Evidence suggests that interleukin-10 (IL-10) deficiency exacerbates inflammation and worsens the outcome of brain ischemia. In view of the critical role of the single nucleotide polymorphic sites -1082 (A/G) and -819 (C/T) in the promoter region of the IL-10 gene, we hypothesized that they are associated with cerebral infarction morbidity in the Chinese Han population. We genotyped these allelic gene polymorphisms by amplification refractory mutation system-polymerase chain reaction methods in 181 patients with cerebral infarction (cerebral infarction group) and 115 healthy subjects (control group). We identified significant differences in genotype distribution and allele frequency of the IL-10-1082 A/G allele between cerebral infarction and control groups (χ (2) = 6.643, P = 0.010). The IL-10-1082 A allele frequency was significantly higher in the cerebral infarction group (92.3%) than in the control group (86.1%) (P = 0.015). Moreover, cerebral infarction risk of the AA genotype was 2-fold higher than with the AG genotype (OR = 2.031, 95%CI: 1.134-3.637). In addition, AA genotype together with hypertension was the independent risk factor of cerebral infarction (OR = 2.073, 95%CI: 1.278-3.364). No statistical difference in genotype distribution or allele frequency of IL-10-819 C/T was found between cerebral infarction and control groups (P > 0.05). These findings suggest that the IL-10-1082 A/G gene polymorphism is involved in cerebral infarction, and increased A allele frequency is closely associated with occurrence of cerebral infarction.

[Effects of Cu2+ stress on DNA polymorphism of genome in foxtail millet of different genotypes].

Cu2+ is an essential element for plant growth, and is one of the major elements in the environment. In order to investigate the physiological characteristics and geno-toxicity effects of foxtail millet (Setaria italica (L) Beauv) under different Cu2+ stress, four genotypes of foxtail millet (Zhaogu, Huangmi, An06, D2-8) from Shanxi, China were cultivated for 30 days in a pot filled with soil of with different mass concentrations of Cu2+ (0, 50, 100, 200, 400 mg.kg-l). Effects of Cu2+ stress on DNA damage of genome in foxtail millet were studied using random amplified polymorphic DNA (RAPD) , and the contents of soluble sugar, proline and MDA were tested. The result showed that the content of soluble sugar had a trend of initial increased followed by decline in all four foxtail millet seedlings in response to the rising Cu2+ concentration, and the maximum value was 50 mg.kg-1. At Cu2 concentrations of 200 mg. kg-1 or more, the soluble sugar content in the four kinds of millet showed an average reduction of 32.44% to 56.5% compared to that of the control group. The result showed that proline synthesis was enhanced at low concentrations (less than 50 mg.kg-1) , but inhibited at high concentrations (more than 100 mg.kg-1), and the contents of MDA in the four genotypes of foxtail millet were significantly increased compared with the control group (P <0. 05). The changes occurring in random amplified polymorphic DNA profiles of the four genotypes of foxtail millet following Cu' treatment included loss of normal bands, appearance of new bands and variation in band intensity compared to the plantlet without treatment, showing that Cu2+ significantly affected the stability of the genomic DNA in the cells of millet seedlings. Additionally, the effect of DNA polymorphism changes was dose-dependent with the Cu2+ concentration. The different genotypes of millet showed different response in the physiological and genetic damage under Cu2+ stress. The change of DNA polymorphism using RAPD technique could be used as the biomarkers to find genotoxic effects of Cu2+.

An Arabidopsis mutant atcsr-2 exhibits high cadmium stress sensitivity involved in the restriction of H2S emission.

The gene AtCSR encodes peptidyl-prolyl cis/trans isomerases (PPIases) that accelerate energetically unfavorable cis/trans isomerization of the peptide bond preceding proline production. In our studies, we found that AtCSR was associated with cadmium (Cd)-sensitive response in Arabidopsis. Our results show that AtCSR expression was triggered by Cd-stress in wild type Arabidopsis. The expression of some genes responsible for Cd(2+) transportation into vacuoles was induced, and the expression of the iron-regulated transporter 1 (IRT1) related to Cd(2+) absorption from the environment was not induced in wild type with Cd(2+) treatment. The expression of Cd-transportation related genes was not in response to Cd-stress, whereas IRT expression increased dramatically in atcsr-2 with Cd(2+) treatment. The expression of glutathione 1 (GSH1) was consistent with GSH being much lower in atcsr-2 in comparison with the wild type with Cd(2+) treatment. Additionally, malondialdehyde (MDA), hydrogen peroxide, and Cd(2+) contents, and activities of some antioxidative enzymes, differed between the wild type and atcsr-2. Hydrogen sulfide (H(2)S) has been confirmed as the third gas-transmitter over recent years. The findings revealed that the expression pattern of H(2)S-releasing related genes and that of Cd-induced chelation and transportation genes matched well in the wild type and atcsr-2, and H(2)S could regulate the expression of the Cd-induced genes and alleviate Cd-triggered toxicity. Finally, one possible suggestion was given: down-regulation of atcsr-2, depending on H(2)S gas-transmitter not only weakened Cd(2+) chelation, but also reduced Cd(2+) transportation into vacuoles, as well as enhancing the Cd(2+) assimilation, thus rendering atcsr-2 mutant sensitive to Cd-stress.

[The effects of adipose-derived stem cells transplantation on the expression of TGF-ß1 in rat brain after cerebral ischemia].

AIM: To explore the mechanisms of adipose-derived stem cells (ADSCs) transplanting induced angiogenesis in rat brain after focal cerebral ischemia. METHODS: 108 male adult Sprague-Dawley rats were randomly assigned into 4 groups: sham-operated group, middle cerebral artery occlusion (MCAO) group, vehicle group and MCAO+ADSCs-treated group. Rat's focal cerebral ischemia model was established by right middle cerebral artery occlusion (MCAO) with modified Longa's method. ADSCs were pre-labeled by CFSE before the transplantation into the rat brain. At 1 d after MCAO, 30 µL cell suspension which contained 1×10(6); ADSCs was injected into the lateral ventricle of MCAO+ADSCs-treated rats, and the same dose of PBS was given to the rats of vehicle group as control. At 4 d, 7 d and 14 d after MCAO, rats were decapitated to detect the TGF-ß1 expression in the infarct area. RESULTS: The expression of TGF-ß1 in brain tissues in MCAO+ADSCs-treated group was significantly higher than MCAO group and vehicle group at 4 d, 7 d and 14 d after MCAO, respectively. After transplantation into MCAO rats, ADSCs could survive and express TGF-ß1 in the ischemic brain. CONCLUSION: These data suggest that ADSCs transplantation can promote revascularization in cerebral ischemic rats, partly by promoting TGF-ß1 expression in the brain.

[Effects of adipose-derived stem cells transplantation on the neuronal apoptosis and the expression of Bcl-2 and caspase-12 in the brain post focal cerebral ischemia in rats].

AIM: To investigate the effects of adipose-derived stem cells (ADSCs) transplantation on neuronal apoptosis in the brain after focal cerebral ischemia in rats. METHODS: 72 male adult Sprague-Dawley rats were randomly divided into 4 groups: Sham-operated group , Middle cerebral artery occlusion (MCAO) group, Vehicle group and ADSC-treated group (n=18). MCAO model was established with the modified Longa's method. One day after right MCAO, 30 µL of cell suspension containing 1×10(6); cells were injected into the lateral ventricle of ADSC-treated group and the same dose of PBS was given to the vehicle group. At 4 d, 7 d and 14 d after MCAO, the apoptosis of neuron was detected by terminal deoxynucleotidyl transferase-mediated DNA nick-end labeling (TUNEL) method and the expression of Bcl-2 and caspase-12 in ischemic region was detected by immunohistochemistry and RT-PCR. RESULTS: TUNEL-positive cells in ischemic region of ADSC-treated group were less than that in MCAO group and Vehicle group at 4 d, 7 d and 14 d post MCAO (P<0.05). Compared with MCAO group and Vehicle group, the expression of Bcl-2 significantly up-regulated while caspase-12 expression significantly decreased in ADSC-treated group at any time point post MCAO (P<0.05). CONCLUSION: The transplantation of ADSCs can reduce neuronal apoptosis of rats with cerebral ischemic injury partly by promoting the expression of Bcl-2 which participates in apoptotic signals after mitochondrial damage and inhibiting the expression of caspase-12 which mediates endoplasmic reticulum (ER) stress-induced apoptosis.

[Interleukin-10 inhibits the expression of interferon regulatory factor-1 with anoxia-reoxygenation in vitro].

AIM: to explore the effects of IL-10 on the expression of interferon regulatory factor-1 (IRF-1) after anoxia-reoxygenation in vitro. METHODS: brain microvascular endothelial cells (BMEC) and neutrophil (PMN) were co-cultured exposed to eight hours of anoxia followed by two hours of reoxygenation to establish anoxia-reoxygenation model in vitro. The co-cultured BMEC and PMN were randomly divided into the following groups: normal group, anoxia-reoxygenation group, 1 microg/L IL-10 treatment group, 10 microg/L IL-10 treatment group, and 30 microg/L IL-10 treatment group. The expression of IRF-1 was detected by RT-PCR and Western blot method. RESULTS: the expression of IRF-1 was significantly upregulated after anoxia-reoxygenation. IL-10 inhibited the expression of IRF-1 in a dose-dependent manner compared with anoxia-reoxygenation group, and the peak of inhibitive effects occurred at the dose of 30 µg/L. CONCLUSION: IL-10 can inhibit the expression of interferon regulatory factor-1 with anoxia-reoxygenation in vitro.

[The effects of adipose-derived stem cell transplantation on the expression of IL-10 and TNF-alpha after cerebral ischaemia in rats].

AIM: To explore the effects of adipose-derived stem cell (ADSC) transplantation on the expression of IL-10 amd TNF-alpha after cerebral ischaemia in Middle cerebral artery occlusion (MCAO) rats. METHODS: 72 male adult Sprague-Dawley rats were randomly divided into 4 groups: sham group, MCAO group, Vehicle group and ADSC group (n=18). Rat's cerebral ischemia model was established by MCAO with Longa's method. ADSC were labeled by DAPI before transplantation. One day after MCAO, 30 microL of cell suspension containing 1 x 10(6); ADSCs were injected into the lateral ventricle of ADSC group and the same dose of PBS was given to the Vehicle group. At day 4, day 7 and day 14 after MCAO, the rats were decapitated to detect the expression of IL-10 and TNF-alpha in ischaemic rat's brain by ELISA, immunohistochemistry and RT-PCR. RESULTS: Compared with sham group, the expression of IL-10 and TNF-alpha significantly up-regulated at 4 d, 7 d of MCAO group(P<0.05). There was no statistical difference of IL-10 and TNF-alpha expression between MCAO and vehicle group ant any time point(P>0.05). Compared with Vehicle group, the expression of IL-10 significantly up-regulated while TNF-alpha expression significantly decreased of ADSC-treated group at any timepoint post MCAO(P<0.05). CONCLUSION: The transplantation of ADSC could up-regulate the expression of IL-10 and down-regulate the expression of TNF-alpha in MCAO rat's brain, which might contribute to its protective role upon cerebral ischaemia.

[Inhibition effect and its mechanism of nobiletin on proliferation of lung cancer cells].

OBJECTIVE: To investigate inhibition effect and its mechanism of Nobiletin on the proliferation of lung cancer cells in vivo and in vitro. METHODS: Human lung adenocarcinoma cell line A549 was treated with various dose of Nobiletin, and cell proliferation was tested by MTT assay. DNA damage of A549 cells was detected by comet assay. Expression of Bcl-2 and Bax proteins were analyzed by Western blot assay. Lewis lung carcinoma (LLC) models were established in C57BL/6 mice and treated with Nobiletin once a day for 12 d, and then the inhibitory rate of tumor growth was measured. Cellular apoptosis was determined using TUNEL assay. The protein levels of Bax, Bcl-2 and Caspase-9 in LLC tissues were determined by immunohistochemistry. RESULTS: The treatment of Nobiletin resulted in significant inhibition of A549 cells proliferation in a dose-dependent manner. Comet assay revealed that DNA damage increased along with the increase of Nobiletin dosage. Western blot analysis showed that A549 cells pretreated with Nobiletin had Bcl-2 protein expression decreased and Bax protein expression increased. The inhibitory rates of lung carcinoma were 43.70%, 27.59% and 20.14% in lewis mice treated with high (300 mg/kg), middle (200 mg/kg), and low (100 mg/kg) dosage of Nobiletin (P<0.01). With the treatment of Nobiletin, the expression of Bax and Caspase-9 proteins increased, and the expression of Bcl-2 proteins decreased. CONCLUSION: Nobiletin has certain inhibitory effects on the proliferation of lung cancer cells both in vivo and in vitro. The mechanism may be related to up-regulation of Bax and Caspase-9 and down-regulation of Bcl-2.

[Clinical observation on therapeutic effect of electroacupuncture at Quchi (LI 11) for treatment of essential hypertension].

OBJECTIVE: To observe the therapeutic effect of electroacupuncture (EA) at Quchi (LI 11) on blood pressure and blood plasma catecholamines in the patient of essential hypertension and to investigate the mechanism. METHODS: Sixty cases of essential hypertension were randomly divided into an EA group (n=30) and a control group (n=30). In the EA group, bilateral Quchi (LI 11) were selected; and in the control group, western medicine Nicardipine was taken. The variation of blood pressure and blood plasma catecholamines were examined before and after the treatment. RESULTS: (1) After treatment, there were significant reduction in the levels of systolic blood pressure and diastole blood pressure in both groups (P < 0.01); (2) After treatment, significant reduction in levels of adrenaline and noradrenaline were also found in both groups (P < 0.01), however, no significant differences in the level of dopamine were observed in both groups (P > 0.05); (3) The effective rate of 66.7% in the EA group was similar to that of 70.0% in the control group (P > 0.05). CONCLUSION: Both EA at Quchi (LI 11) and western medicine are able to beneficially regulate blood pressure of patients with essential hypertension through adjusting blood plasma catecholamines.

[Effects of adipose-derived stem cell transplantation on the angiogenesis and the expression of bFGF and VEGF in the brain post focal cerebral ischemia in rats].

AIM: To investigate the effects of adipose-derived stem cell (ADSC) transplantation on the angiogenesis in the brain post focal cerebral ischemia in rats. METHODS: 72 male adult Sprague-Dawley rats were randomly divided into 4 groups: sham-operated group, middle cerebral artery occlusion (MCAO) group, vehicle group and MCAO+ADSC-treated group (n=18). A permenant focal cerebral ischemia model was established with the modified Longa's method. ADSC were labeled by DAPI before transplantation. One day after right MCAO, 30 muL of cell suspension containing 1x10(6) cells were injected into the lateral ventricle of MCAO+ADSC-treated group and the same dose of PBS was given to the vehicle group. On D4, D7 and D14 after MCAO, the rats were killed to detect the regeneration of microvessel and the expression of bFGF and VEGF in ischemic region by immunohistochemistry and RT-PCR. RESULTS: A lot of microvessel proliferate in the injured cortex reached peak in 2 weeks. The microvessel density in the brain tissues of rats treated with ADSC was higher than that in MCAO group and vehicle group (P<0.01). The expression of bFGF and VEGF in the brain tissues of MCAO+ADSC-treated group was higher than that in MCAO group and vehicle group on D4, D7 and D14 post MCAO. CONCLUSION: The transplantation of ADSC can promote the revascularization of cerebral ischemia in rats partly by enhancing bFGF and VEGF synthesis in brain.

[Effects of bone marrow-derived mesenchymal stem cells transplantation on recovery of neurological functions and expression of synaptophysin in focal cerebral infarction in rats].

AIM: To investigate the effects of bone marrow-derived mesenchymal stem cells (BMSC) transplantation on the recovery of neurological functions and the expression of synaptophysin in focal cerebral infarction in rats. METHODS: 72 male adult Sprague-Dawley rats were randomly divided into 4 groups (18 in each group): shamoperated group, middle cerebral artery occlusion (MCAO) roup,vehicle group and MCAO+BMSC-treated group. A permanent focal cerebral ischemia model was established using modified Longa's method. BMSC was labeled by DAPI before the transplantation. One day after right middle cerebral artery occlusion(MCAO), 1 x 10(6) cells were injected into the lateral ventricle of rats in BMSCs-treated group and the same dose of PBS was given to the rats in vehicle group. Before sacrificed and at 4 d, 7 d and 14 d after MCAO, the neurological functions were tested by balance beam, rota-rod and screen prehensile and the synaptophysin was detected by reverse transcriptase-polymerase chain reaction (RT-PCR) and immunohistochemical method. RESULTS: DAPI stained positive cells were observed around the cerebral infarcted area in the BMSC-treated group. Compared with the MCAO group and the vehicle group,the neurological functions in BMSC-treated group were better on 7 d and 14 d after MCAO (P<0.05 or P<0.01), and the synaptophysin around the cerebral infarcted area was significantly upregulated on 4 d, 7 d and 14 d after MCAO (all P<0.01). CONCLUSION: BMSC transplantation can improve the neurological functions by upregulating the expression of synaptophysin after MCAO in rats.