Aequorivita vitellina sp. nov. and Aequorivita xiaoshiensis sp. nov., isolated from marine sediment.

Two Gram-stain-negative, strictly aerobic, chemoheterotrophic, short-rod-shaped and non-motile strains, forming yellow colonies and designated F47161T and F64183T, were isolated from marine sediment of Xiaoshi Island, Wei Hai, PR China. Strain F47161T grew at 15-37 °C (optimally at 30 °C) and pH 6.0-9.0 (optimally at pH 7.5) and in the presence of 1-9 % (w/v) NaCl (optimally at 3 %). Strain F64183T grew at 10-37 °C (optimally at 30 °C) and pH 6.0-8.5 (optimally at pH 7.0) and in the presence of 1-8 % (w/v) NaCl (optimally at 3 %). The results of phylogenetic analysis based on 16S rRNA gene sequences indicated that F47161T and F64183T were related to members of the genus Aequorivita. The strains shared 97.4 % 16S rRNA gene sequence similarity to each other. F47161T and F64183T shared highest 16S rRNA gene sequence similarity to Aequorivita sinensis JCM 19789T, and the values were 97.5 % and 98.4 %, respectively. The predominant cellular fatty acids of both F64183T and F47161T were iso-C15 : 0 and iso-C17 : 0 3-OH, but the predominant fatty acids of F47161T also included anteiso-C15 : 0. The sole respiratory quinone of F47161T and F64183T was menaquinone 6 (MK-6), consistent with that observed for all related strains. Between F47161T and F64183T, the average nucleotide identity (ANI) and digital DNA-DNA hybridisation (dDDH) values were 75.8 % and 20.5 %, respectively, and between the novel isolates (F47161T and F64183T) and A. sinensis JCM 19789T they were 76.0 % and 94.2 % and 20.6 % and 57.1 %, respectively. The genomic DNA G+C contents of F47161T and F64183T was 37.3 % and 34.5 %, respectively. The polar lipid profiles of F47161T and F64183T contained phosphatidylethanolamine, two aminolipids, one glycolipid, one phosphoglycolipid and two unidentified polar lipids. Differential phenotypic and genotypic characteristics of the two strains indicated that the two strains should be classified as representing two novel species of the genus Aequorivita, for which the names Aequorivita vitellina sp. nov. and Aequorivita xiaoshiensis sp. nov. are proposed. The type strains are F47161T (=MCCC 1H00509T=KCTC 92017T) and F64183T (=MCCC 1H00507T=KCTC 92016T), respectively.

Cerina litoralis gen. nov., sp. nov., a novel potential polysaccharide degrading bacterium of the family Flavobacteriaceae, isolated from marine sediment.

The Gram-strain-negative, facultative anaerobic, chemoheterotrophic, short-rod-shaped, non-motile, forming yellow colonies strain, designated F89T, was isolated from marine sediment of Xiaoshi Island, Weihai. Strain F89T grew at 15-37 °C (optimally at 28 °C), at pH 6.0-8.5 (optimally at pH 7.0) and in the presence of 1-5% (w/v) NaCl. Phylogenetic analysis based on 16S rRNA gene sequence showed that strain F89T was related to the family Flavobacteriaceae. F89T had highest 16S rRNA gene sequence similarity to Maribacter cobaltidurans MCCC 1K03318T (93.3%). The predominant cellular fatty acids of F89T were iso-C15:0, iso-C15:0 G and Summed Feature 3. The main respiratory quinone of F89T was menaquinone 6 (MK-6), consistent with that observed for all related strains. The polar lipid profile of strain F89T contained phosphatidylethanolamine, two aminolipids and three unidentified polar lipids. The genomic DNA G + C content of strain F89T was 42.7%. Strain F89T encoded 121 glycoside hydrolases and was a potential polysaccharide degrading bacterium. Differential phenotypic and genotypic characteristics of the strain showed that F89T should be classified as a novel genus in Flavobacteriaceae, for which the name Cerina litoralis is proposed. The type strain is F89T (= MCCC 1H00510T = KCTC 92203T).

Winogradskyella immobilis sp. nov., an Alginate-Hydrolyzing Bacterium Isolated from the Brown Algae Saccharina japonica.

A novel bacterium, designated E313T, was isolated from brown algae Saccharina japonica in Weihai, China. The strain is a Gram-stain-negative, non-flagellated, non-gliding, aerobic, rod-shaped bacterium that grows optimally at 28 °C with pH levels between 7.0 and 7.5 and in the presence of 2-3% (w/v) NaCl. Phylogenetic analyses based on its 16S rRNA gene sequence placed the strain within the monophyletic cluster of the genus Winogradskyella, exhibiting the highest similarity to Winogradskyella wandonensis KCTC 32579T (96.8%). Genome comparison of strain E313T with W. wandonensis KCTC 32579T and W. thalassocola KCTC 12221T revealed average nucleotide identity (ANI) values of 74.2% and 74.8%, and DNA-DNA hybridization (dDDH) values of 19.0% and 19.5%, respectively, lower than prokaryotic species delineation values. The strain E313T could hydrolyze alginate. A total of 123 carbohydrate-active enzymes were annotated according to the CAZy database. Especially, one oligo-alginate lyase and one poly(ß-D-mannuronate) lyase were identified in the genome of strain E313T. Strain E313T possessed MK-6 quinone and iso-C15:0, iso-C15:1 G, iso-C17:0 3-OH, and iso-C15:0 3-OH as main fatty acids. Its major polar lipids were phosphatidylethanolamine (PE), one unidentified aminolipid, and two unknown lipids. Thus, based on phylogenetic, physiological, and chemotaxonomic analyses, we propose a novel species of the genus Winogradskyella, named Winogradskyella immobilis sp. nov., with E313T (= MCCC 1H00506T = KCTC 82731T) as the type strain.

Aestuariirhabdus haliotis sp. nov., isolated from abalone viscera and emended description of the genus Aestuariirhabdus.

Two novel strains, Z083T and Z084, were isolated from the viscera of abalone, Haliotis discus hannai, sampled in Weihai, PR China. The phenotypic, chemotaxonomic and genomic characteristics of the two strains were studied. The average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values between the two strains were 99.8 and 98.9 %, respectively, suggesting that the two strains belonged to the same species. The 16S rRNA gene sequence analysis showed 99.8 % similarity between the two strains, while the genome analysis indicated that they were not from one clonal origin. Phylogenetic analysis of 16S rRNA gene sequences showed the two strains belonged to the genus Aestuariirhabdus and Aestuariirhabdus litorea JCM 32043T was the closest strain (97.5 %). Genomic analysis, including calculations of ANI, dDDH, amino acid identity (AAI) and percentage of conserved proteins (POCP), between Z083T, Z084 and A. litorea JCM 32043T clearly separated those two strains from A. litorea JCM 32043T as the values were below the thresholds for species delineation. The genome size of strains Z083T and Z084 were approximately 4.16 and 4.23 Mbp, respectively, and the DNA G+C contents of both strains were 51.8 mol%. According to the phenotypic, chemotaxonomic and phylogenetic characterizations and the results of genome analysis, Z083T and Z084 could be identified as belonging to a novel species of the genus Aestuariirhabdus, for which the name Aestuariirhabdus haliotis sp. nov., is proposed, with Z083T (=MCCC 1H00501T=KCTC 92006T) as the type strain.

Risk probability and influencing factors of stroke in followed-up hypertension patients.

OBJECTIVE: To explore the risk probability and main influencing factors of stroke in followed-up hypertension patients through the analysis of long-term followed-up cohort data. METHODS: The method of followed-up observation cohort was used to collect the information of 168,417 followed-up hypertension patients from 2002 to 2020 in Jiading District in Shanghai. Kaplan-Meier method was used to analyze the risk probability of stroke complications in long-term followed-up HTN patients, and the influencing factors were analyzed by Cox proportional risk model. RESULTS: Among 168,417 followed-up hypertension patients, 11,143 cases had suffered stroke, and the cumulative incidence rate of stroke was 6.62% (male was 6.87%, female was 6.37%). With the extension of the hypertension years, the cumulative risk probability of stroke in HTN patients would continue to increase and the interval was not equidistant. The total cumulative risk probability of stroke in HTN patients was 78.9% (male was 91.0%, female was 70.7%). During the period of hypertension, the risk occurring probability of stroke was not fixed, but fluctuating. There were 4 onset peaks, which were in 8 years (probability was 4.2%), 15 years (probability was 14.0%), 22 years (probability was 6.0%) and 26 years (probability was 13.9%). The highest risk probability of male patients was in 26 years (probability was 23.1%), and the second peak was in 15 years (probability was 15.6%). The highest risk probability of female patients was in 15 years (probability was 12.9%), and the second peak was in 26 years (probability was 8.7%). The risk probability of different gender, BP grade and BMI was different, the male were at higher risk than the female, stage 3 HTN was higher than stage 2 and stage 1 HTN, obese people and underweight people were at higher risk than those who have normal weight. The main factors closely related to the occurrence of stroke complications were age (RR = 2.917, p < 0.001), body mass index (RR = 1.654, p < 0.001), family history of stroke (RR = 1.386, p < 0.001) and blood pressure grade (RR = 1.148, p < 0.001). CONCLUSION: The risk probability of stroke among hypertension patients was high in followed-up hypertension patients (total 78.9%, male 91.0%, female 70.7%), and would continue to increase disproportionately during period of hypertension (4 different onset peaks). With the persistence of hypertension, the risk probability of stroke would increase continuously. Multivariate Cox regression analysis showed that male patients, patients with HBP, abnormal BMI and positive family history were main factors closely related to the occurrence of stroke complications.

Cutaneous Immunoprofiles of Three Spotted Fever Group Rickettsia Cases.

Spotted fever group rickettsia (SFGR) can cause mild to fatal illness. The early interaction between the host and rickettsia in skin is largely unknown, and the pathogenesis of severe rickettsiosis remains an important topic. A surveillance of SFGR infection by PCR of blood and skin biopsy specimens followed by sequencing and immunohistochemical (IHC) detection was performed on patients with a recent tick bite between 2013 and 2016. Humoral and cutaneous immunoprofiles were evaluated in different SFGR cases by serum cytokine and chemokine detection, skin IHC staining, and transcriptome sequencing (RNA-seq). A total of 111 SFGR cases were identified, including 79 "Candidatus Rickettsia tarasevichiae," 22 Rickettsia raoultii, 8 Rickettsia sibirica, and 2 Rickettsia heilongjiangensis cases. The sensitivity to detect SFGR in skin biopsy specimens (9/24, 37.5%) was significantly higher than that in blood samples (105/2,671, 3.9%) (P < 0.05). As early as 1 day after the tick bite, rickettsiae could be detected in the skin. R. sibirica infection was more severe than "Ca Rickettsia" and R. raoultii infections. Increased levels of serum interleukin-18 (IL-18), IP10, and monokine induced by gamma interferon (MIG) and decreased levels of IL-2 were observed in febrile patients infected with R. sibirica compared to those infected with "Ca Rickettsia." RNA-seq and IHC staining could not discriminate between SFGR-infected and uninfected tick bite skin lesions. However, the type I interferon (IFN) response was differently expressed between R. sibirica and R. raoultii infections at the cutaneous interface. It is concluded that skin biopsy specimens were more reliable for the detection of SFGR infection in human patients although the immunoprofile may be complicated by immunomodulators induced by the tick bite.

Drastic induction of MMP-7 by cortisol in the human amnion: implications for membrane rupture at parturition.

Preterm premature rupture of fetal membranes precedes 30-40% of preterm births. Activation of matrix metalloproteases (MMPs) is the one of the major causes of extracellular matrix (ECM) degradation in membrane rupture. Increased cortisol, regenerated by 11ß-hydroxysteroid dehydrogenase 1 in the amnion at parturition, is known to participate in a number of parturition-pertinent events. However, whether cortisol has a role in the regulation of MMPs in the membranes is not known. Here, we addressed this issue using human amnion tissue, the most tensile layer of the membranes. RNA-sequencing revealed that cortisol induced MMP7 expression dramatically in amnion fibroblasts, which was confirmed by real-time quantitative RT-PCR and Western blotting analysis in cortisol-treated amnion explants and fibroblasts. Measurement of collagen IV α5 chain (COL4A5), a substrate for MMP-7, showed that cortisol reduced its extracellular abundance, which was blocked by an antibody against MMP-7. Moreover, increased MMP-7 but decreased COL4A5 abundance was observed in the amnion tissue following labor-initiated spontaneous rupture of membranes. Mechanistic studies showed that cortisol increased the phosphorylation of c-Jun and the expression of c-Fos, the 2 major components of activated protein 1 (AP-1), respectively. The knocking down of c-Fos or c-Jun significantly attenuated the induction of MMP7 expression by cortisol. Chromatin immunoprecipitation assays showed that cortisol stimulated the enrichment of c-Fos and c-Jun at the AP-1 binding site in the MMP7 promoter. The data suggest that induction of MMP7 by cortisol via AP-1 may be a contributing factor to ECM degradation in membrane rupture at parturition.-Wang, L.-Y., Wang, W.-S., Wang, Y.-W., Lu, J.-W., Lu, Y., Zhang, C.-Y., Li, W.-J., Sun, K., Ying, H. Drastic induction of MMP-7 by cortisol in the human amnion: implications for membrane rupture at parturition.

Involvement of serum amyloid A1 in the rupture of fetal membranes through induction of collagen I degradation.

The de novo synthesis of serum amyloid A1 (SAA1) is augmented in human fetal membranes at parturition. However, its role in parturition remains largely unknown. Here, we investigated whether SAA1 was involved in the rupture of fetal membranes, a crucial event in parturition accompanied with extensive degradation of collagens. Results showed that SAA1 decreased both intracellular and extracellular COL1A1 and COL1A2 abundance, the two subunits of collagen I, without affecting their mRNA levels in human amnion fibroblasts. These reductions were completely blocked only with inhibition of both matrix metalloproteases (MMPs) and autophagy. Consistently, SAA1 increased MMP-2/9 abundance and the markers for autophagic activation including autophagy related (ATG) 7 (ATG7) and the microtubule-associated protein light chain 3 ß (LC3B) II/I ratio with the formation of LC3 punctas and autophagic vacuoles in the fibroblasts. Moreover, the autophagic degradation of COL1A1/COL1A2 and activation of MMP-2/9 by SAA1 were blocked by inhibitors for the toll-like receptors 2/4 (TLR2/4) or NF-κB. Finally, reciprocal corresponding changes of SAA1 and collagen I were observed in the amnion following spontaneous rupture of membranes (ROM) at parturition. Conclusively, SAA1 may participate in membrane rupture at parturition by degradating collagen I via both autophagic and MMP pathways. These effects of SAA1 appear to be mediated by the TLR2/4 receptors and the NF-κB pathway.

Transmission dynamics of Ebola virus disease and intervention effectiveness in Sierra Leone.

Sierra Leone is the most severely affected country by an unprecedented outbreak of Ebola virus disease (EVD) in West Africa. Although successfully contained, the transmission dynamics of EVD and the impact of interventions in the country remain unclear. We established a database of confirmed and suspected EVD cases from May 2014 to September 2015 in Sierra Leone and mapped the spatiotemporal distribution of cases at the chiefdom level. A Poisson transmission model revealed that the transmissibility at the chiefdom level, estimated as the average number of secondary infections caused by a patient per week, was reduced by 43% [95% confidence interval (CI): 30%, 52%] after October 2014, when the strategic plan of the United Nations Mission for Emergency Ebola Response was initiated, and by 65% (95% CI: 57%, 71%) after the end of December 2014, when 100% case isolation and safe burials were essentially achieved, both compared with before October 2014. Population density, proximity to Ebola treatment centers, cropland coverage, and atmospheric temperature were associated with EVD transmission. The household secondary attack rate (SAR) was estimated to be 0.059 (95% CI: 0.050, 0.070) for the overall outbreak. The household SAR was reduced by 82%, from 0.093 to 0.017, after the nationwide campaign to achieve 100% case isolation and safe burials had been conducted. This study provides a complete overview of the transmission dynamics of the 2014-2015 EVD outbreak in Sierra Leone at both chiefdom and household levels. The interventions implemented in Sierra Leone seem effective in containing the epidemic, particularly in interrupting household transmission.

Human Babesiosis Caused by a Babesia crassa-Like Pathogen: A Case Series.

Background: Human babesiosis is an emerging health problem in China. Methods: Babesia were identified in ticks, sheep, and humans in northeastern China using polymerase chain reaction (PCR) followed by genetic sequencing. We enrolled residents who experienced a viral-like illness after recent tick bite or were healthy residents. We defined a case using the definition for babesiosis developed by the US Centers for Disease Control and Prevention. Results: A Babesia crassa-like agent was identified in Ixodes persulcatus and Haemaphysalis concinna ticks using PCR followed by sequencing. The agent was characterized through phylogenetic analyses of the 18S rRNA gene, the ß-tubulin gene, and the internal transcribed spacer region. We tested sheep as a possible reservoir and found that 1.1% were infected with the B. crassa-like agent. We screened 1125 human participants following tick bites using B. crassa-specific PCR and identified 31 confirmed and 27 suspected cases. All the patients were previously healthy except for 1 with an ovarian tumor. Headache (74%), nausea or vomiting (52%), and fever (48%) were the most common clinical manifestations of confirmed cases. Six of 10 cases remained PCR positive for B. crassa-like infection 9 months after initial diagnosis. Asymptomatic infections were detected in 7.5% of 160 local residents. Conclusions: We identified B. crassa-like infection in people in northeastern China that caused mild to moderate symptoms. The possibility of more severe disease in immunocompromised patients and of transmission through the blood supply due to asymptomatic infections justifies further investigation of this reported infection.

Borrelia miyamotoi Infections in Humans and Ticks, Northeastern China.

We conducted an investigation of Borrelia miyamotoi infections in humans and ticks in northeastern China. Of 984 patients reporting recent tick bites, 14 (1.4%) were found to be infected with B. miyamotoi by PCR and genomic sequencing. The 14 patients had nonspecific febrile manifestations, including fever, headache, anorexia, asthenia, and arthralgia. Rash, eschar, and regional lymphadenopathy were each observed in 1 patient. Four (28.6%) patients were hospitalized because of severe disease. B. miyamotoi was detected in 3.0% (19/627) of Ixodes persulcatus, 1 (2.8%) of 36 Haemaphysalis concinna, and none of 29 Dermacentor silvarum ticks. Phylogenetic analyses based on sequences of a nearly entire 16s rRNA gene, a partial flagellin gene, and the glycerophosphodiester phosphodiesterase gene revealed that B. miyamotoi identified in patients and ticks were clustered in the group of the Siberian type. These findings indicate that B. miyamotoi is endemic in northeastern China and its public health significance deserves further investigation.

Candidatus Rickettsia tarasevichiae Infection in Eastern Central China: A Case Series.

BACKGROUND: Human infection with Candidatus Rickettsia tarasevichiae (CRT) was first reported in northeastern China in 2012. OBJECTIVE: To describe the clinical spectrum and laboratory findings of patients infected with CRT in eastern central China. DESIGN: Case series. SETTING: A sentinel hospital for severe fever with thrombocytopenia syndrome (SFTS) in eastern central China in 2014. PARTICIPANTS: Hospitalized patients with SFTS-like illness. MEASUREMENTS: Molecular and serologic tests were performed to diagnose CRT infection. Data about clinical manifestations and laboratory findings were retrieved from medical records. RESULTS: 56 of 733 assessed patients had CRT based on polymerase chain reaction and sequencing. All patients presented with nonspecific manifestations, including fever (96%), malaise (88%), myalgia (57%), cough (25%), and dizziness (14%). Only 2 patients had rash. Further, 16% had eschar, 29% had lymphadenopathy, 100% had gastrointestinal symptoms, 34% had neurologic symptoms, 43% had hemorrhagic manifestations, and 23% had signs of plasma leakage. Thrombocytopenia was observed in 70%, leukopenia in 59%; lymphopenia in 45%; and elevated levels of lactate dehydrogenase in 82%, aspartate aminotransferase in 70%, alanine aminotransferase in 54%, and creatinine kinase in 46%. Co-infection with SFTS virus was documented in 66% patients, and 8 of the 56 patients died. LIMITATIONS: Patients with CRT were not treated for infection because they were retrospectively identified. This was not a population-based study, and the results cannot be generalized to all patients with CRT. CONCLUSION: Candidatus R tarasevichiae infection should be considered in the differential diagnosis of febrile patients with SFTS-like illness in endemic areas. PRIMARY FUNDING SOURCE: National Natural Science Foundation of China.

Integrated Exploitation of the Structural Diversity Space of Chemotherapy Drugs to Selectively Inhibit HER2 T798M Mutant in Lung Cancer.

An acquired T798M gatekeeper mutation in human epidermal growth factor receptor 2 (HER2) kinase can cause drug resistance to anti-HER2 chemotherapy drugs in lung cancer. Previously, the reversible pan-kinase inhibitor staurosporine has been found to selectively inhibit the HER2 T798M mutant over wild-type kinase, suggesting that the staurosporine scaffold is potentially to develop mutant-selective inhibitors. Here, we systematically evaluated the chemical space of staurosporine scaffold-based compounds in response to HER2 T798M mutation at structural, energetic and molecular levels by using an integrated analysis strategy. With this strategy, we were able to identify several novel wild-type sparing inhibitors with high or moderate selectivity, which are comparable to or even better than that of the parent compound staurosporine. Molecular modeling and structural analysis revealed that noncovalent contacts can form between the side chain of mutated residue Met798 and selective inhibitor ligands, which may improve the favorable interaction energy between the kinase and inhibitor and reduce the unfavorable desolvation penalty upon the kinase-inhibitor binding.

Renal AML with inferior vena cava thrombus treated by workbench surgery and autotransplantation.

Renal angiomyolipoma may sometimes require surgical intervention. In this paper, we present a case of renal angiomyolipoma which infiltrated the sinus and extended into the inferior vena cava. He was successfully treated with a combined approach of laparoscopic radical nephrectomy, extracorporeal workbench tumor resection, and autotransplantation. Three months postoperatively, no evidence of tumor recurrence or presence of thrombus in the inferior vena cava was noted. Our experience represents the successful application of a combined nephron-sparing approach in the management of angiomyolipoma with extension into a major blood vessel.

DNAJA1­knockout alleviates heat stroke­induced endothelial barrier disruption via improving thermal tolerance and suppressing the MLCK­MLC signaling pathway.

Endothelial barrier disruption plays a key role in the pathophysiology of heat stroke (HS). Knockout of DNAJA1 (DNAJA1­KO) is thought to be protective against HS based on a genome­wide CRISPR­Cas9 screen experiment. The present study aimed to illustrate the function of DNAJA1­KO against HS in human umbilical vein endothelial cells. DNAJA1­KO cells were infected using a lentivirus to investigate the role of DNAJA1­KO in HS­induced endothelial barrier disruption. It was shown that DNAJA1­KO could ameliorate decreased cell viability and increased cell injury, according to the results of Cell Counting Kit­8 and lactate dehydrogenase assays. Moreover, HS­induced endothelial cell apoptosis was inhibited by DNAJA1­KO, as indicated by Annexin V­FITC/PI staining and cleaved­caspase­3 expression using flow cytometry and western blotting, respectively. Furthermore, the endothelial barrier function, as measured by transepithelial electrical resistance and FITC­Dextran, was sustained during HS. DNAJA1­KO was not found to have a significant effect on the expression and distribution of cell junction proteins under normal conditions without HS. However, DNAJA1­KO could effectively protect the HS­induced decrease in the expression and distribution of cell junction proteins, including zonula occludens­1, claudin­5, junctional adhesion molecule A and occludin. A total of 4,394 proteins were identified using proteomic analysis, of which 102 differentially expressed proteins (DEPs) were activated in HS­induced wild­type cells and inhibited by DNAJA1­KO. DEPs were investigated by enrichment analysis, which demonstrated significant enrichment in the 'calcium signaling pathway' and associations with vascular­barrier regulation. Furthermore, the 'myosin light­chain kinase (MLCK)­MLC signaling pathway' was proven to be activated by HS and inhibited by DNAJA1­KO, as expected. Moreover, DNAJA1­KO mice and a HS mouse model were established to demonstrate the protective effects on endothelial barrier in vivo. In conclusion, the results of the present study suggested that DNAJA1­KO alleviates HS­induced endothelial barrier disruption by improving thermal tolerance and suppressing the MLCK­MLC signaling pathway.

[RIN1 expression in hepatocellular cancer and the affection on prognosis and tumor invasion ability].

OBJECTIVE: To test the expression of RIN1 in hepatocellular cancer (HCC) and study its clinicopathological significance and mechanism. METHODS: RIN1 mRNA in 36 HCC tissues was analyzed using real-time PCR (RT-PCR). The expression of RIN1 was examined by immunohistochemistry (IHC) in 110 HCC specimens. The relationship between the protein expression and prognosis was analyzed. Transwell was used to test invasion ability of HCC cell lines which were transfected with the expression vector pEGFP-N1-RIN1. RESULTS: RIN1 mRNA expression levels was much lower in tumor tissues than that in their corresponding non-cancerous tissues (χ(2) = 7.430, P = 0.026). RIN1 protein was lowly expressed in liver cancer samples (69.1%) and correlated with poor survival (6.46%) (χ(2) = 13.808, P < 0.05). Transwell assays show that RIN1 overexpression can inhibit invasion ability of HepG2 cells (t = 8.975 and 9.522, both P < 0.05). RIN1 expression and ABL2 and E calcium protein were positively correlated (r = 0.898 and 0.912, P < 0.05), and negatively correlated with MMP-9 (r = -0.933, P = 0.002). CONCLUSIONS: RIN1 expression was down-regulated in HCC and low expression of RIN1 foreshows poor prognosis of HCC patients. RIN1 overexpression can inhibit invasion ability of HepG2 cells.

Modifying the amino acids in conformational motion pathway of the α-amylase of Geobacillus stearothermophilus improved its activity and stability.

Amino acids along the conformational motion pathway of the enzyme molecule correlated to its flexibility and rigidity. To enhance the enzyme activity and thermal stability, the motion pathway of Geobacillus stearothermophilus α-amylase has been identified and molecularly modified by using the neural relational inference model and deep learning tool. The significant differences in substrate specificity, enzymatic kinetics, optimal temperature, and thermal stability were observed among the mutants with modified amino acids along the pathway. Mutants especially the P44E demonstrated enhanced hydrolytic activity and catalytic efficiency (kcat/KM) than the wild-type enzyme to 95.0% and 93.8% respectively, with the optimum temperature increased to 90°C. This mutation from proline to glutamic acid has increased the number and the radius of the bottleneck of the channels, which might facilitate transporting large starch substrates into the enzyme. The mutation could also optimize the hydrogen bonding network of the catalytic center, and diminish the spatial hindering to the substrate entry and exit from the catalytic center.

Changes of Serum Trace Elements, AFP, CEA, SF, T3, T4 and IGF-II in Different Periods of Rat Liver Cancer.

OBJECTIVE: Based on liver cancer model built in SD rats, the contents of trace elements (Cu, Fe, Zn, Ca and Mg), AFP, CEA, SF, TH and IGF-II in serum were measured at different stages to explore the molecular changes during the rat liver cancer development. METHODS: The SD rat liver cancer model was built by using diethylnitrosamine (DENA) as the mutagen. During 16 weeks of DENA gavage, blood samples were taken in the 14th, 28th, 56th, 77th, 105th and 112th days respectively after the first day of gavage with DENA, then the contents of five trace elements (Cu, Fe, Zn, Ca and Mg), T3, T4, IGF-II, AFP, CEA and SF in serum were determined. RESULTS: During the development of the rat liver cancer, in the test group, the Cu content significantly increased in serum, while the contents of Fe, Zn and Ca significantly decreased. The content of Mg showed no significant change. AFP and CEA of the test group showed same expression level with the control group; while the content of SF was lower than that of the control group when cancerization appeared. T3 and T4 increased at the first stage and then went down, and the content of IGF-II was always high. CONCLUSION: Cu, Fe, Zn, Ca, T3, T4, SF and IGF-II are closely related to the development of liver cancer. The changes of their contents in the development of cancer could enlighten the researches on cancer pathogenesis and prevention.

The near-infrared dye IR-61 restores erectile function in a streptozotocin-induced diabetes model via mitochondrial protection.

This study aimed to evaluate the therapeutic effect of IR-61, a novel mitochondrial heptamethine cyanine dye with antioxidant effects, on diabetes mellitus-induced erectile dysfunction (DMED). Eight-week-old male Sprague-Dawley rats were intraperitoneally injected with streptozotocin (STZ) to induce type 1 diabetes. Eight weeks after STZ injection, all rats were divided into three groups: the control group, DM group, and DM + IR-61 group. In the DM + IR-61 group, the rats were administered IR-61 (1.6 mg kg-1) twice a week by intravenous injection. At week 13, erectile function was evaluated by determining the ratio of the maximal intracavernous pressure to mean arterial pressure, and the penises were then harvested for fluorescent imaging, transmission electron microscopy, histological examinations, and Western blot analysis. Whole-body imaging suggested that IR-61 was highly accumulated in the penis after intravenous injection. IR-61 treatment significantly improved the maximal ICP of diabetic rats. Additionally, IR-61 ameliorated diabetes-induced inflammation, apoptosis, and phenotypic transition of corpus cavernosum smooth muscle cells (CCSMCs) in penile tissue. IR-61 also attenuated mitochondrial damage, reduced reactive oxygen species production in the corpus cavernosum and upregulated sirtuin1 (SIRT1), sirtuin3 (SIRT3), nuclear factor (erythroid-derived 2)-like 2 (Nrf2), and heme oxygenase expression in penile tissue. In conclusion, IR-61 represents a potential therapeutic option for DMED by protecting the mitochondria of CCSMCs, which may be mediated by activation of the SIRT1, SIRT3, and Nrf2 pathways.

[Effect of siRNA-mediated beta-catenin gene on Wnt signal pathway in lung adenocarcinoma A549 cell].

OBJECTIVE: To study the effects of siRNA-mediated beta-catenin down-regulation on Wnt/beta-catenin signal transduction pathway in lung adenocarcinoma A549 cells. METHODS: A549 cell was divided into three groups: PGCsil-CTNNB1-siRNA group (transfection with beta-catenin interference plasmids), negative control group (transfection with negative control plasmids) and non-transfection group (blank control). For each group, real-time PCR was employed to detect the expression of beta-catenin and cyclin D1 (a target gene of Wnt/beta-catenin pathway). A MTT cell proliferation assay was performed to study the proliferating capacity. Flow cytometry was used for cell cycle analysis. Clone formation and Millicell chamber experiment were performed to evaluate the clone formation and migration capacities of cells. RESULTS: The expression of beta-catenin and cyclin D1 in PGCsil-CTNNB1-siRNA cell decreased at the mRNA level. It was lower than negative control (0.002+/-0.001 vs 0.023+/-0.002, P<0.01; 0.005+/-0.002 vs 0.040+/-0.020, P<0.05) and blank control groups (beta-catenin mRNA: 0.021+/-0.003, P<0.01; cyclin D1 mRNA: 0.042+/-0.004, P<0.05). Also the proliferating capacity at Days 5-7 was inhibited in comparison with negative control and blank control groups (P<0.05, P<0.01). The cell-doubling time (58.1 h) was markedly longer than those of negative control group (37.9 h, P<0.05) and blank control group (34.2 h, P<0.05). The number of cells in G0-G1 phrase increased in PGCsil-CTNNB1-siRNA group (86.4%+/-2.6%) in comparison with negative control (73.8%+/-0.9%, P<0.01) and blank control groups (75.8%+/-1.5%, P<0.01). In PGCsil-CTNNB1-siRNA group, the clone formation ratio (31.6%+/-7.7%) was lower than negative control (46.9%+/-7.3%, P<0.05) and blank control groups (44.2%+/-2.5%, P<0.05). And the number of cells (16.0+/-3.8) passing through the membrane of Millicell chamber was smaller than negative control (32.7+/-3.1, P<0.01) and blank control groups (33.0+/-2.7, P<0.01). CONCLUSIONS: Knocking down beta-catenin not only inhibits the Wnt/beta-catenin signal transduction pathway in lung adenocarcinoma A549 cells but also decreases cell proliferation, clone formation and migration capacity. Thus it may become a novel target for lung cancer therapy.