Association analysis of the gut microbiota in predicting outcomes for patients with acute ischemic stroke and H-type hypertension.

Introduction: H-type hypertension (HHTN) is a subtype of hypertension that tends to worsen the prognosis of acute ischemic stroke (AIS). Recent studies have highlighted the vital role of gut microbiota in both hypertension and AIS, but there is little available data on the relationship between gut microbiota and the progression of AIS patients with HHTN. In this study, we investigated the microbial signature of AIS patients with HHTN and identified characteristic bacteria as biomarkers for predicting prognosis. Methods: AIS patients with HHTN (n = 150) and without HHTN (n = 50) were enrolled. All patients received a modified Rankin Scale (mRS) assessment at 3 months after discharge. Fecal samples were collected from the participants upon admission, including 150 AIS patients with HHTN, 50 AIS patients with non-HHTN, and 90 healthy subjects with HHTN. These samples were analyzed using 16S rRNA sequencing to characterize the bacterial taxa, predict functions, and conduct correlation analysis between specific taxa and clinical features. Results: Our results showed that the composition of the gut microbiota in HHTN patients differed significantly from that in non-HHTN patients. The abundance of the genera Bacteroides, Escherichia-Shigella, Lactobacillus, Bifidobacterium, and Prevotella in AIS patients with HHTN was significantly increased compared to AIS patients without HHTN, while the genus Streptococcus, Faecalibacterium, and Klebsiella were significantly decreased. Moreover, Bacteroides, Lactobacillus, Bifidobacterium, and Klebsiella in AIS patients with HHTN were more abundant than healthy subjects with HHTN, while Escherichia-Shigella, Blautia, and Faecalibacterium were less abundant. Moreover, the genera Butyricicoccus, Rothia, and Family_XIII_UCG-001 were negatively connected with the NIHSS score, and the genera Butyricicoccus and Rothia were observed to be negatively associated with the mRS score. The genera Butyricicoccus, Romboutsia, and Terrisporobacter were associated with a poor prognosis, whereas the increase in Butyricimonas and Odoribacter was correlated with good outcomes. Generated by eight genera and clinical indexes, the area under the curve (AUC) value of the receiver operating characteristic (ROC) curve achieved 0.739 to effectively predict the prognosis of AIS patients with HHTN. Conclusion: These findings revealed the microbial signature of AIS patients with HHTN and further provided potential microbial biomarkers for the clinical diagnosis of AIS patients with HHTN.

Activation of STIM1/Orai1­mediated SOCE in sepsis­induced myocardial depression.

Unbalanced Ca2+ homeostasis serves an essential role in the occurrence and development of septic myocardial injury. However, the mechanism of Ca2+ homeostasis in septic myocardial depression is poorly understood due to the complexity of Ca2+ transporters in excitable cells. It was therefore hypothesized that cardiac dysfunction, myocardial injury and cardiac apoptosis in septic myocardial depression are associated with elevated intracellular Ca2+ concentrations caused by stromal interaction molecule 1 (STIM1)/Orai calcium release­activated calcium modulator 1 (Orai1)­mediated store­operated Ca2+ entry (SOCE). A septic myocardial depression model was established using the cecal ligation and puncture operation (CLP) in mice and was simulated in H9C2 cells via lipopolysaccharide (LPS) stimulation. Cardiac function, myocardial injury, cardiac apoptosis and the expression levels of Bax, Bcl­2, STIM1 and Orai1 were quantified in vivo at 6, 12 and 24 h. Changes in the intracellular Ca2+ concentration, SOCE and the distribution of STIM1 were assessed in vitro within 6 h. The morphological changes of heart tissue were observed by hematoxylin­eosin staining. Myocardial cellular apoptosis was determined by TUNEL method. The expression of Bax, Bcl­2, STIM1 and Orai1 were visualized by western blot. Cytosolic calcium concentration and SOCE were evaluated by confocal microscopy. The results demonstrated that cardiac contractile function was significantly reduced at 6 h and morphological changes in cardiac tissues, as well as the myocardial apoptosis rate, were markedly increased at 6, 12 and 24 h following CLP. mRNA and protein expression levels of Bax/Bcl­2 were significantly enhanced at 6 and 12 h and glycosylation of Orai1 in the myocardium of septic mice was significantly increased at 6 h following CLP. The intracellular Ca2+ concentration, SOCE, was significantly increased at 1­2 h and the clustering and distribution of STIM1 were markedly changed in H9C2 cells at 1 and 2 h. These findings suggested that myocardial dysfunction, cardiac injury and myocardial depression may be related to increased intracellular Ca2+ concentration resulting from STIM1/Orai1­mediated SOCE, which may provide a potential method to alleviate septic myocardial depression.

Plasma Concentration After the First Hemoperfusion has a High Predictive Value in Medium Level Acute Paraquat-Poisoned Patients.

BACKGROUND: Paraquat ( PQ) is very poisonous to humans and animals and there is no effective clinical antidote . The efficacy of hemoperfusion (HP) treatment for PQ poisoning remains controversial. To explore new ways to predict the prognosis of patients with acute PQ poisoning and assist in the development of better hemopurification treatment strategies. METHODS: The clinical data of patients who were intoxicated with PQ through contact were diagnosed with PQ poisoning by high-performance liquid chromatography. Samples were collected by the Emergency Intensive Care Unit of the First Affiliated Hospital of Wenzhou Medical University from January 2012 to November 2016. Based on the prognosis, the patients were grouped into survival and death groups. Comparisons of the differences in the clinical indexes were performed, including the initial concentration of PQ at admission, PQ concentration after first HP, the number of HP cartridges used for the first hemoperfusion, whether HP was combined with continuous renal replacement therapy, and the number of concurrent organ injuries between the 2 groups. In addition, data were analyzed using multivariate logistic regression models and receiver operating characteristic curves. Moreover, prognostic factors in patients with acute PQ poisoning were analyzed. RESULTS: Overall, 128 patients with acute PQ poisoning were enrolled in this study. The median plasma PQ concentrations of the patients at admission were 21 and 834 ng/mL (range: 50-1,099,118 ng/mL). The multiple logistic regression model revealed that the initial concentration of PQ and the PQ concentration after the first perfusion were independent risk factors for death in patients with acute PQ poisoning. The PQ concentration in the survival group after the first HP was <516 ng/mL and was mainly distributed at approximately 100 ng/mL. The percentage of patients whose concentration after the first HP was <516 ng/mL in the death group was only 19%. CONCLUSIONS: The initial plasma PQ concentration after admission and PQ concentration after the first HP are risk factors for death in patients with acute PQ poisoning. Moreover, PQ concentration after the first HP had a high predictive value for death. When the initial plasma PQ concentration after admission ranges from 50 ng/mL to 5000 ng/mL, the rapid reduction in plasma PQ concentration after HP treatment could improve the prognosis of patients with acute PQ poisoning.

Rosiglitazone Inhibits Activation of Hepatic Stellate Cells via Up-Regulating Micro-RNA-124-3p to Alleviate Hepatic Fibrosis.

BACKGROUND: The activation of hepatic stellate cells (HSCs) is involved in hepatic fibrogenesis and is regulated by the decreased expression of peroxisome proliferator-activated receptor γ (PPARγ). Rosiglitazone (RGZ) is a highly potent agonist of PPARγ. AIMS: To clarify molecular regulatory mechanism of RGZ in the activation of HSCs in hepatic fibrosis. METHODS: A mouse model of hepatic fibrosis was established by carbon tetrachloride with or without RGZ intervention. A vector carrying pcDNA-HOTAIR was constructed and injected into a mouse model. HSCs were isolated from liver tissue and activated by transforming growth factor-ß. The expression of miR-124-3p, HOTAIR, Col1A1, α-SMA, and PPARγ mRNAs was measured by quantitative real-time PCR. The level of PPARγ was measured by Western blotting. The interaction between HOTAIR and PPARγ was assessed by RNA immunoprecipitation (RIP) and RNA pull-down. The target gene of miR-124-3p was determined by luciferase reporter assay and RNA interference approaches. RESULTS: The expression of Col1A1 and α-SMA was reduced after RGZ intervention. Different expressions of HOTAIR and miR-124-3p were observed in liver tissue and HSCs. The luciferase reporter assay and RNA interference approaches indicated that miR-124-3p negatively regulated HOTAIR expression. RIP and RNA pull-down results revealed that PPARγ was interacted by HOTAIR. The therapeutic effect of RGZ on hepatic fibrosis was reversed by overexpression of HOTAIR. CONCLUSIONS: RGZ inhibits the activation of HSCs by up-regulating miR-124-3p. The silencing of HOTAIR by miR-124-3p in HSC activation provided the foundation to understand interactions of ncRNAs and potential treatment target in hepatic fibrosis.

Comparison of bone marrow-vs. adipose tissue-derived mesenchymal stem cells for attenuating liver fibrosis.

Mesenchymal stem cell (MSC) therapy has emerged as a potential novel method of treating liver fibrosis. To date, bone marrow-derived MSCs (BM-MSCs) and adipose tissue-derived MSCs (AD-MSCs) have not been analyzed with respect to their ability to combat liver fibrosis. The present study aimed to compare the capabilities of BM-MSCs and AD-MSCs in the treatment of liver fibrosis. BM-MSCs and AD-MSCs were taken from male Sprague-Dawley rats and cultured. Hepatic stellate cells (HSCs) were co-cultured with either BM-MSCs or AD-MSCs, and the effects of BM-MSCs or AD-MSCs on the proliferation, activation and apoptosis of HSCs were determined. The secretion of a selected group of cytokines by BM-MSCs and AD-MSCs was measured using enzyme-linked immunosorbent assays. Using a CCl4-induced liver fibrosis animal model, the anti-inflammatory and anti-fibrotic effects of BM-MSCs or AD-MSCs against liver fibrosis in vivo were evaluated. The morphological examination and analysis of specific surface markers confirmed the successful preparation of BM-MSCs and AD-MSCs. Furthermore, the proliferation, activation and apoptosis of HSCs were significantly inhibited by BM-MSCs and AD-MSCs, with statistically greater reductions achieved by AD-MSCs compared with BM-MSCs. Direct comparison of the secretion of selected cytokines by BM-MSCs and AD-MSCs revealed that significantly higher levels of nerve growth factor and transforming growth factor-ß1 were secreted in the AD-MSC culture medium, whereas levels of vascular endothelial growth factor and interleukin-10 did not differ significantly between AD-MSCs and BM-MSCs. In vivo studies using a CCl4-induced liver fibrosis model demonstrated that inflammatory activity and fibrosis staging scores were significantly lower in the MSC-treated groups compared with controls. Although AD-MSCs improved anti-inflammatory and anti-fibrotic effects compared with BM-MSCs, these differences were not significant. Thus, the current study demonstrated that BM-MSCs and AD-MSCs are similarly effective at attenuating liver fibrosis by inhibiting the activation and proliferation of HSCs, as well as promoting the apoptosis of HSCs.

Ethyl pyruvate reverses development of Pseudomonas aeruginosa pneumonia during sepsis-induced immunosuppression.

Sepsis is characterized by an innate immune response and the following immune dysfunction which can increase the emergence of secondary infections. Ethyl pyruvate (EP) has multiple immunoregulation functions in several serious illnesses, such as burn injury, severe sepsis and acute respiratory syndrome. However, little data was shown the effect of EP administration on immunosuppression post-CLP and the following secondary infection. The cecal ligation and puncture (CLP) followed by the induction of Pseudomonas aeruginosa (PA) was used as a clinically relevant two-hit model of sepsis. We assessed the survival rate, lung damage and lung bacterial clearance in vehicle or EP treatment group to demonstrate the lung response to Pseudomonas aeruginosa of septic mice. Then cytokines including lung IL-6, IL-1ß, IL-10 and plasma HMGB1, apoptosis of splenic immune cells and Foxp3 level on regulatory T cells (Tregs) were studied to demonstrate the mechanisms of EP administration on two-hit mice. We found that the susceptibility of septic mice to Secondary Pseudomonas aeruginosa pneumonia could be down-regulated by ethyl pyruvate treatment and the protective effects of EP may via decreasing lung IL-10 and plasma HMGB1 expression, inhibiting the function of Tregs and relieving the apoptosis of splenic immune cells. The "immune paralysis" post-sepsis still remains a rigorous challenge for curing sepsis, our study may aid in the development of new therapeutic strategies to this problem.

[Analyses on relevant factors of the prognosis of patients with acute organophosphate poisoning].

OBJECTIVE: To find out a method which can assess the prognosis of patients with Acute Organophosphate Poisoning objectively and increase the successful ratio of treatment by investigating relevant factors on the prognosis of the patients with Acute Organophosphate Poisoning. METHODS: We retrospected 116 patients with Acute Organophosphate Poisoning who were treated in our hospital's emergency room from April 2006 to March 2014. According to the outcome of patients, we distributed the patients to death group and survival group, compared the clinic data and using multivariate analysis with Logistic regression to prognosis factors. RESULTS: 116 cases of acute organophosphate poisoning patients died in 23 cases, improved in 93 cases. Death group patients' APACHE-II score are higher than whose in the survival group (P < 0.05). Compared with the survival group, patients' body temperature, blood pressure, pH, GCS index were lower in the death group (P < 0.05) and Cr, WBC, ALT, AST, CK-MB, blood glucose, blood lactic acid, heart rate were higher in the death group (P < 0.05), there were significant difference between two groups with statistical.Low blood pressure, lower GCS score, hyperglycemia and high white blood cell count, were independent risk factors of poor prognosis, and hypotension was maximum value of all the factor (OR = 54.22). CONCLUSION: APACHE II prognostic scoring system can be accurately response, vital signs, white blood cell count, pH, serum creatinine, GCS score and serum sodium value which in this system may be associated with prognosis. To evaluate the severity and prognosis of illness Blood glucose, ALT, AST, CK-MB's rising also has certain value.

[Hemoperfusion alleviated paraquat-induced kidney inflammation injury of rabbit].

OBJECTIVE: To investigate the effect of hemoperfusion on paraquat-Induced kidney inflammation injury of rabbit and the mechanism of it. METHODS: 60 male rabbits were randomly divided into 4 groups, the normal control group (n=6, the rabbits were given NS by gavage) , blank control group (n=18, he rabbits were given 2 hours hemoperfusion once within 1 hour after given NS by gavage), paraquat poisoning group (n=18, the rabbits were given 50 mg/kg 20% paraquat solution by gavage) , hemoperfusion treatment group (n=18, the rabbits were given 2 hours hemoperfusion once within 1 hour after 20% paraquat solution espoused). The last 3 groups were divided into 3 observation time groups (1, 3, 7 day), contained 6 rabbits each group. On days 1, 3, 7 all groups rabbits were anesthetized and sacrificed, and their kidney tissues collected. The levels of NF-κB mRNA by RT-PCR, and the expression of NF-κB protein was measured by Western blotting,The expression levels of TNF-α, IL-6, iNOS measured by chemical colorimetric method to to observe inflammatory injury. RESULTS: Compared with the normal control group rabbits, there were no changes in the TNF-α, IL-6, iNOS, NF-κB mRNA and protein of blank control group (P>0.05), while the expression of TNF-α, IL-6, NF-κB mRNA and protein in the kidney tissue of PQ group and were significantly increased (P<0.05). The pathological results of kidney tissues were no abnormalities onnormal control group and blank control group. CONCLUSION: HP significantly increase resistance to PQ-induced inflammation injury in the rabbit kidney and exert a protective effect on PQ-induced kidney injury.

[The protective effect of bone marrow mesenchymal stem cell on lung injury induced by vibrio vulnificus sepsis].

OBJECTIVE: To discuss the protective effect of bone marrow mesenchymal stem cell (BMSC) on lung injury induced by vibrio vulnificus sepsis and its mechanism. METHODS: BMSCs were isolated by whole bone marrow adherent culture from mouse. Male ICR mice were randomly divided into normal saline control group (NS group), normal saline + BMSC control group (NSB group), vibrio vulnificus sepsis group (VV group), vibrio vulnificus sepsis + BMSC group (VVB group) according to random number table, with 40 mice in each group. Sepsis mouse model was reproduced by injecting vibrio vulnificus (1 × 107 cfu/mL) 5 mL/kg through the left side peritoneal cavity, and caudal intravenous injection of BMSC (4 × 105 cfu/mL) 5 mL/kg for intervention after model reproduction. Ten mice in each group were sacrificed at 6, 12, 24 or 48 hours after injecting vibiro vulnificus, and their lung tissues were harvested. The lung wet/dry (W/D) ratio was calculated. The expression of nuclear factor-ΚBp65 (NF-ΚBp65) in nucleus was measured by Western Blot. The levels of tumor necrosis factor-α (TNF-α) and interleukins (IL-1ß, IL-6) in lung tissue were detected by enzyme-linked immunosorbent assay (ELISA). The pathological changes in lung tissue were observed after hematoxylin-eosin (HE) staining and uranyl acetate-lead citrate staining. RESULTS: After vibrio vulnificus injection, lung W/D ratio, the expression of NF-ΚBp65 in nucleus, and the levels of TNF-α, IL-1ß, IL-6 in the lung tissues were significantly increased in VV group compared with those in NS group at all the time points, and peaked at 12 hours. Compared with the VV group, the VVB group had significantly decreased levels of lung W/D ratio, NF-ΚBp65 expression, and the levels of TNF-α, IL-1ß, IL-6, with significant differences at all the time points [VV group vs. NS group at 12 hours: lung W/D ratio 7.22 ± 0.03 vs. 5.21 ± 0.02, NF-ΚBp65 expression (glay scale) 1.86 ± 0.74 vs. 0.75 ± 0.07, TNF-α (ng/L) 433.24 ± 3.23 vs. 106.57 ± 1.21, IL-1ß (ng/L) 35.64 ± 0.15 vs. 10.64 ± 0.48, IL-6 (ng/L) 58.84 ± 0.55 vs. 17.69 ± 1.35, all P<0.05; VVB group vs. VV group at 12 hours: lung W/D ratio 6.49 ± 0.06 vs. 7.22 ± 0.03, NF-ΚBp65 expression (A value) 1.16 ± 0.08 vs. 1.86 ± 0.74, TNF-α (ng/L) 357.22 ± 3.25 vs. 433.24 ± 3.23, IL-1ß (ng/L) 27.77 ± 0.59 vs. 35.64 ± 0.15, IL-6 (ng/L) 38.6 8 ± 1.29 vs. 58.84 ± 0.55, all P<0.05]. There were no significant differences in above indexes between NS group and NSB group. In the NS and NSB groups pathological changes were not obvious under light microscopy, in the VV group lung tissue hyperemia and edema was significant, the edema fluid, red blood cells and inflammatory cells also could be seen, and in the VVB group lung damage that mentioned above could be alleviated. In the NS and NSB groups epithelial cell structure of type I and type II was completed, and the changes were not obvious under the transmission electron microscopy. In the VV group the alveolar walls were damaged significantly, with type I epithelial cell cytoplasm swelling, bubbling and rupture, with type II epithelial cells visible cytoplasm decrease, cavitation, addiction to osmium lamellar corpuscle emptying, lysosome hyperplasia, microvilli reduction, and in the VVB group the above damage was alleviated. CONCLUSIONS: Vibrio vulnificus sepsis can cause acute lung damage and edema, and BMSC can down regulate inflammatory cytokines, reduce lung injury caused by vibrio vulnificus sepsis.

Mitochondrial tRNA mutations are associated with maternally inherited hypertension in two Han Chinese pedigrees.

We report here the clinical, genetic, molecular, and biochemical evaluations in two Han Chinese families with maternally inherited hypertension. Fourteen of 20 adult matrilineal relatives of these families exhibited a wide range of severity in hypertension, while none of offspring of affected fathers had hypertension. The age-at-onset of hypertension in matrilineal relatives varied from 37 years to 83 years, with an average of 55 and 66 years, respectively. Mutational analysis of their mitochondrial genomes identified the m.4353T>C mutation in the tRNA, in conjunction with the known m.593C>T mutation in the tRNA(Phe) and m.5553C>T mutation in the tRNA(Trp). Northern analysis revealed that m.4353T>C, m.593C>T and m.5553C>T mutations caused ∼66%, 65%, and 12% reductions in the steady-state level of tRNA(Gln), tRNA(Phe) and tRNA(Trp), respectively. An in vivo protein labeling analysis showed ∼35% reduction in the rate of mitochondrial translation in cells carrying these tRNA mutations. Impaired mitochondrial translation is apparently a primary contributor to the reduced rates of overall respiratory capacity, malate/glutamate-promoted respiration, succinate/glycerol-3-phosphate-promoted respiration, or N,N,N',N'-tetramethyl-p-phenylenediamine/ascorbate-promoted respiration and the increasing level of reactive oxygen species in the cells carrying these mtDNA mutations. These data demonstrate that mitochondrial dysfunction caused by mitochondrial tRNA mutations is associated with essential hypertension in these families.

The tRNAMet 4435A>G mutation in the mitochondrial haplogroup G2a1 is responsible for maternally inherited hypertension in a Chinese pedigree.

Mutations in mitochondrial DNA (mtDNA) have been associated with hypertension in several pedigrees with maternal inheritance. However, the pathophysiology of maternally inherited hypertension remains poorly understood. We reported here clinical, genetic evaluations and molecular analysis of mtDNA in a three-generation Han Chinese family with essential hypertension. Eight of 17 matrilineal relatives exhibited a wide range of severity in essential hypertension, whereas none of the offsprings of the affected father had hypertension. The age-at-onset of hypertension in the maternal kindred varied from 31 to 65 years, with an average of 52 years. Sequence analysis of mtDNA in this pedigree identified the known homoplasmic 4435A>G mutation, which is located at immediately 3' end to the anticodon, corresponding to the conventional position 37 of tRNA(Met), and 41 variants belonging to the Asian haplogroup G2a1. In contrast, the 4435A>G mutation occurred among mtDNA haplogroups B5a, D, M7a2 and J. The adenine (A37) at this position of tRNA(Met) is extraordinarily conserved from bacteria to human mitochondria. This modified A37 was shown to contribute to the high fidelity of codon recognition, structural formation and stabilization of functional tRNAs. However, 41 other mtDNA variants in this pedigree were the known polymorphisms. The occurrence of the 4435A>G mutation in two genetically unrelated families affected by hypertension indicates that this mutation is involved in hypertension. Our present investigations further supported our previous findings that the 4435A>G mutation acted as an inherited risk factor for the development of hypertension. Our findings will be helpful for counseling families of maternally inherited hypertension.