Changes in peripheral blood TBNK lymphocyte subsets and their association with acute exacerbation of chronic obstructive pulmonary disease.

OBJECTIVE: This study aimed to determine the changes in peripheral blood TBNK lymphocyte subsets in patients with acute exacerbation of chronic obstructive pulmonary disease (AECOPD) and their relationship with the pathogenesis of AECOPD. METHODS: A cross-sectional study of 1252 hospitalized patients in Zhejiang Hospital was conducted. There were 162 patients in the AECOPD group and 1090 in the non-chronic obstructive pulmonary disease (COPD) group. The proportions of peripheral blood T helper cells, cytotoxic T cells, total B cells, total natural killer (NK) cells, and total T cells in the two groups were determined, and the CD4/CD8 ratio was calculated. RESULTS: The proportions of men and total natural killer cells, and the mean age were significantly higher in the AECOPD group than in the non-COPD group. The T helper cell, total T cell, and CD4/CD8 ratios were significantly decreased in the AECOPD group. A multivariate logistic regression analysis showed that male sex, age, the total T cell ratio, and the CD4/CD8 ratio were significantly associated with the incidence of AECOPD. CONCLUSION: Cellular immune dysfunction in patients with AECOPD causes a decrease in total T lymphocytes and the CD4/CD8 ratio, which may be involved in the pathogenesis of AECOPD.

Overexpression of an Antioxidant Enzyme APX1 in cpr5 Mutant Restores its Pleiotropic Growth Phenotype.

Breeding crops with enhanced immunity is an effective strategy to reduce yield loss caused by pathogens. The constitutive expresser of pathogenesis-related genes (cpr5) mutant shows enhanced pathogen resistance but retarded growth; thus, it restricts the application of cpr5 in breeding crops with disease resistance. Reactive oxygen species (ROS) play important roles in plant growth and defense. In this study, we determined that the cpr5 mutant exhibited excessive ROS accumulation. However, the mutation of respiratory burst oxidase homolog D (RBOHD), a reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase responsible for the production of ROS signaling in plant immunity, did not suppress excessive ROS levels in cpr5. Furthermore, the cpr5 mutant showed low levels of ascorbate peroxidase 1 (APX1), an important cytosolic ROS-scavenging enzyme. APX1 overexpression in the cpr5 background removed excessive ROS and restored the pleiotropic growth phenotype. Notably, APX1 overexpression did not reduce the resistance of cpr5 mutant to virulent strain Pseudomonas syringae pv. tomato (Pst) DC3000 and avirulent strain Pst DC3000 (avrRpt2). These results suggest that the removal of excessive ROS by APX1 overexpression restored the cpr5 growth phenotype while conserving pathogen resistance. Hence, our study provides a theoretical and empirical basis for utilizing CPR5 in the breeding of crops with disease resistance by effective oxidative stress management via APX1 expression.

Ascorbate peroxidase 1 allows monitoring of cytosolic accumulation of effector-triggered reactive oxygen species using a luminol-based assay.

Biphasic production of reactive oxygen species (ROS) has been observed in plants treated with avirulent bacterial strains. The first transient peak corresponds to pattern-triggered immunity (PTI)-ROS, whereas the second long-lasting peak corresponds to effector-triggered immunity (ETI)-ROS. PTI-ROS are produced in the apoplast by plasma membrane-localized NADPH oxidases, and the recognition of an avirulent effector increases the PTI-ROS regulatory module, leading to ETI-ROS accumulation in the apoplast. However, how apoplastic ETI-ROS signaling is relayed to the cytosol is still unknown. Here, we found that in the absence of cytosolic ascorbate peroxidase 1 (APX1), the second phase of ETI-ROS accumulation was undetectable in Arabidopsis (Arabidopsis thaliana) using luminol-based assays. In addition to being a scavenger of cytosolic H2O2, we discovered that APX1 served as a catalyst in this chemiluminescence ROS assay by employing luminol as an electron donor. A horseradish peroxidase (HRP)-mimicking APX1 mutation (APX1W41F) further enhanced its catalytic activity toward luminol, whereas an HRP-dead APX1 mutation (APX1R38H) reduced its luminol oxidation activity. The cytosolic localization of APX1 implies that ETI-ROS might accumulate in the cytosol. When ROS were detected using a fluorescent dye, green fluorescence was observed in the cytosol 6 h after infiltration with an avirulent bacterial strain. Collectively, these results indicate that ETI-ROS eventually accumulate in the cytosol, and cytosolic APX1 catalyzes luminol oxidation and allows monitoring of the kinetics of ETI-ROS in the cytosol. Our study provides important insights into the spatial dynamics of ROS accumulation in plant immunity.

The receptor-like cytosolic kinase RIPK activates NADP-malic enzyme 2 to generate NADPH for fueling ROS production.

Reactive oxygen species (ROS) production is a conserved immune response in Arabidopsis primarily mediated by respiratory burst oxidase homolog D (RBOHD), a nicotinamide adenine dinucleotide phosphate (NADPH) oxidase associated with the plasma membrane. A rapid increase in NADPH is necessary to fuel RBOHD proteins and thus maintain ROS production. However, the molecular mechanism by which NADPH is generated to fuel RBOHD remains unclear. In this study, we isolated a new mutant allele of FLAGELLIN-INSENSITIVE 4 (FIN4), which encodes the first enzyme in de novo NAD biosynthesis. fin4 mutants show reduced NADPH levels and impaired ROS production. However, FIN4 and other genes involved in NAD- and NADPH-generating pathways are not highly upregulated upon elicitor treatment, raising a possibility that a cytosolic NADP-linked dehydrogenase might be post-transcriptionally activated to maintain the NADPH supply close to RBOHD. To verify this possibility, we isolated the proteins associated with RPM1-INDUCED PROTEIN KINASE (RIPK), a receptor-like cytoplasmic kinase that regulates broad-spectrum ROS signaling in plant immunity, and identified NADP-malic enzyme 2 (NADP-ME2), an NADPH-generating enzyme. Compared with wild-type plants, nadp-me2 mutants display decreased NADP-ME activity, lower NADPH levels, and reduced ROS production in response to immune elicitors. Furthermore, we found that RIPK can directly phosphorylate NADP-ME2 and enhance its activity in vitro. The phosphorylation of the NADP-ME2 S371 residue contributes to ROS production upon immune elicitor treatment and susceptibility to the necrotrophic bacterium Pectobacterium carotovorum. Collectively, our study suggests that RIPK phosphorylates and activates NADP-ME2 to rapidly increase cytosolic NADPH, thus fueling RBOHD to sustain ROS production in plant immunity.

Sarcomatoid carcinoma of the gallbladder: a case report.

Sarcomatoid carcinoma is a rare type of gallbladder cancer with no specific clinical manifestation. The final diagnosis depends on pathological and immunohistochemical examination. Sarcomatoid carcinoma is characterized by early lymphatic metastasis, rapid progression, a high short-term recurrence rate, and a worse prognosis than adenocarcinoma. This report describes a 60-year-old woman with poorly differentiated adenocarcinoma of the gallbladder. She underwent treatment with chemotherapy and surgery. Palliative surgery was performed for treatment of tumor recurrence in April 2018. Postoperative pathology showed infiltration of poorly differentiated carcinomas, most of which were sarcomatoid. After four cycles of chemotherapy, the disease continued to progress. Anlotinib tablets were given from August 2018 to November 2018 but were then stopped because of gastrointestinal bleeding. The patient died in April 2019. This paper reports the whole process of diagnosis and treatment in this case of gallbladder sarcomatoid carcinoma, thus providing a reference for treatment.

Association Between Body Composition and Frailty in Elder Inpatients.

PURPOSE: The study aimed to investigate the association between body composition and frailty in elder inpatients. PATIENTS AND METHODS: This is a cross-sectional study including 656 elder inpatients (275 females and 381 males) aged ≥65 years, from department of geriatrics of Zhejiang Hospital between January 2018 and March 2019. Sociodemographic, health-related data and anthropometric measurements were evaluated. Body composition was assessed by bioimpedance analysis (BIA), mainly including skeletal muscle mass, body fat mass, total body water, fat-free mass,percent body fat, basal metabolic rate. Frailty was assessed by Clinical Frailty Scale (CFS). Univariate logistic regression was used to analyze the association between body composition and frailty. RESULTS: Frailty was present in 43.9% of the participants. Frail inpatients showed higher waist circumference, body fat mass and percent body fat, non-frail inpatients showed greater upper arm circumference, calf circumference, skeletal muscle mass, total body water, fat-free mass and basal metabolic rate. Subjects with underweight (body mass index (BMI)<18.5 kg/m2; odds ratio (OR), 95% confidence interval (CI)=4.146 (1.286-13.368) P=0.017) and those with high waist circumference (OR 95% CI=1.428 (0.584-3.491) P<0.001), body fat mass (OR, 95% CI=1.143 (0.892-1.315) P<0.001) presented a higher risk of frailty compared to normal subjects. Skeletal muscle mass (OR; 95% CI=0.159 (0.064-0.396) P<0.001) was a protective factor for frailty. CONCLUSION: Frailty in elder Chinese inpatients was characterized by a body composition phenotype with underweight, high waist circumference, low skeletal muscle mass and high body fat mass. Underweight, abdominal obesity and sarcopenic obesity may, therefore, be targets for intervention of frailty.

[Paeoniflorin improves the permeability of cardiac microvascular endothelial cells by regulating Src/vascular endothelial-cadherin pathway].

OBJECTIVE: To investigate the effect and mechanism of paeoniflorin on the permeability of cardiac microvascular endothelial cells (CMECs) in sepsis. METHODS: Primary rat CMECs were isolated and cultured in vitro, and the cells in the logarithmic growth phase were used for experiments. Tetramethylazozolium colorimetry (MTT) was used to screen the safe and effective concentrations of paeoniflorin at 10, 20, and 40 µmol/L. The cells were divided into blank control group, lipopolysaccharide (LPS) group and low, medium and high concentration paeoniflorin pretreatment group. The cells in the blank control group were cultured in complete medium; the cells in the LPS group were challenged with LPS (1 mg/L) in complete medium; and the cells in the paeoniflorin pretreatment groups were pretreated with 10, 20, and 40 µmol/L paeoniflorin at 4 hours before LPS stimulation. The cells in each group were further cultured for 24 hours after LPS stimulation. The horseradish peroxidase (HRP) method was used to detect the permeability of rat CMECs. The enzyme-linked immunosorbent assay (ELISA) was used to detect the CXC chemokine ligand (CXCL1, CXCL2) levels in the cell supernatant. The real-time fluorescence quantitative reverse transcription-polymerase chain reaction (RT-qPCR) was used to detect the mRNA expressions of CXCL1 and CXCL2 in the cells. Western Blot was used to detect phosphorylated Src (p-Src), vascular endothelial-cadherin (VE-cadherin) and phosphorylated mitogen activated protein kinase (p-MAPK). RESULTS: Compared with the blank control group, the permeability of rat CMECs in the LPS group was significantly increased. The cell permeability was improved to some extent after paeoniflorin pretreatment at different concentrations, and the improvement was most obvious in the 40 µmol/L paeoniflorin group, with statistically significant difference as compared with the LPS group (A value: 1.61±0.07 vs. 2.13±0.06, P < 0.01). ELISA results showed that there were moderate amounts of CXCL1 and CXCL2 in the cell supernatant of rat CMECs in the blank control group. However, the secretion of CXCL1 and CXCL2 in the cell supernatant was increased significantly under the induction of LPS. After pretreatment with paeoniflorin at different concentrations, the secretion of CXCL1 and CXCL2 in the cell supernatant was significantly reduced. The most obvious inhibitory effect on CXCL1 was 40 µmol/L paeoniflorin, and the most obvious inhibition on CXCL2 was 20 µmol/L paeoniflorin, the differences were statistically significant as compared with the LPS group [CXCL1 (ng/L): 337.51±68.04 vs. 829.86±65.06, CXCL2 (ng/L): 4.48±0.11 vs. 9.41±0.70, both P < 0.01]. RT-qPCR results showed that the mRNA expressions of CXCL1 and CXCL2 in the rat CMECs were consistent with the ELISA results. LPS could increase mRNA expressions of CXCL1 and CXCL2 in the rat CMECs, and pretreatment with different concentrations of paeoniflorin could significantly reduce the mRNA expressions of CXCL1 and CXCL2. The 40 µmol/L paeoniflorin had the best inhibitory effect on CXCL1 mRNA expression, and the 20 µmol/L paeoniflorin had the best inhibitory effect on CXCL2 mRNA expression, the differences were statistically significant as compared with the LPS group [CXCL1 mRNA (2-ΔΔCt): 0.543±0.004 vs. 0.812±0.089, CXCL2 mRNA (2-ΔΔCt): 10.52±0.71 vs. 17.68±1.09, both P < 0.01]. Western Blot results showed that moderate amounts of p-Src, VE-cadherin and p-MAPK proteins were expressed in the rat CMECs in the blank control group. After LPS stimulation, the expressions of p-Src and p-MAPK proteins were increased significantly, while the expression of VE-cadherin protein was decreased significantly. After pretreatment with different concentrations of paeoniflorin, the expressions of p-Src and p-MAPK proteins in the cells were decreased to varying degrees, while the expression of VE-cadherin protein was increased, and 40 µmol/L paeoniflorin had the most obvious effect, the differences were statistically significant as compared with the LPS group [p-Src protein (p-Src/GAPDH): 1.02±0.09 vs. 1.29±0.05, p-MAPK proteins (p-MAPK/GAPDH): 0.24±0.02 vs. 0.62±0.02, VE-cadherin protein (VE-cadherin/GAPDH): 0.64±0.03 vs. 0.31±0.02, all P < 0.01]. CONCLUSIONS: Paeoniflorin can regulate the Src/VE-cadherin pathway in CMECs, inhibit the expression and secretion of inflammation-related proteins and chemokines, and improve the cell permeability of CMECs induced by LPS.

Relationship between nutritional status and frailty in hospitalized older patients.

OBJECTIVE: The definition of frailty still lacks quantitative biomarkers. This study aimed to investigate the relationship between nutrition-related biomarkers and frailty in hospitalized older patients. MATERIALS AND METHODS: This is a cross-sectional study including 380 hospitalized older patients. The patients were categorized as nonfrail (n=140), prefrail (n=81), and frail (n=159) by the criteria of frailty phenotype. The nutritional status was assessed using the mini nutritional assessment-short form (MNA-SF), levels of serum transferrin (TNF), prealbumin (PA), total protein (TP), albumin (ALB), retinol-binding protein (RBP), and hemoglobin (Hb). RESULTS: The grip strength, levels of serum TFN, TP, ALB, Hb, and MNA-SF scores all decreased significantly in the order of nonfrail, prefrail, and frail groups (P<0.01). Older ages, more fall incidents, and higher polypharmacy ratio were observed in the frail and prefrail groups than in the nonfrail group (P<0.05). Univariate logistic regression analysis showed that frailty was positively related to age, polypharmacy, fall history, nutritional status, levels of TFN, PA, TP, ALB, RBP, and Hb, but was negatively related to grip strength. Ordinal logistic regression analysis showed that older patients who were well nourished, with higher levels of TFN, TP, and ALB were less likely to develop into frailty. CONCLUSION: Hospitalized older patients with better nutritional status and higher levels of TFN, TP, and ALB were less likely to develop into frailty. These nutrition-related biomarkers may be used for the evaluation of nutritional status and frailty in older patients.

Multiple diabetic complications, as well as impaired physical and mental function, are associated with declining balance function in older persons with diabetes mellitus.

OBJECTIVE: To investigate whether there is a difference in balance function between older persons with and without diabetes mellitus (DM), and to identify whether mediating factors, such as diabetic complications, Instrumental Activities of Daily Living (IADL) score, Mini-Mental State Examination (MMSE) score, as well as hemoglobin A1c (HbA1c), fasting plasma glucose (FPG), serum total cholesterol (TC), triglycerides (TG), and low-density lipoprotein (LDL), are associated with balance function in older persons with DM. METHODS: In this cross-sectional study, a total of 208 older persons were divided into a DM group (n=80) and a control group who did not have DM (n=128). Balance function was evaluated with the Tinetti performance-oriented mobility assessment (POMA), which includes balance and gait subscales. Activities of daily living (ADL), IADL, and the MMSE were also measured. Fall incidents in last 12 months, the use of walking aids, fear of falling, comorbidities, and polypharmacy were recorded. Diabetic complications were recorded, and HbA1c, FPG, TC, TG, and LDL were measured in the patients of the DM group. RESULTS: Fall incidents in last 12 months were higher in the DM group than in the control group (P<0.01). POMA score as well as ADL and IADL scores were lower in the diabetic group than the control group (P<0.05). Within the diabetic group, the POMA score was positively related to the ADL score (odds ratio [OR], 11.7; 95% confidence interval [CI], 3.076-44.497; P<0.01), IADL score (OR, 16.286; 95% CI, 4.793-55.333; P<0.01), and MMSE score (OR, 10.524; 95% CI, 2.764-40.074; P<0.01), but was negatively related to age (OR, 7.707; 95% CI, 2.035-29.185; P<0.01) and diabetic complication (OR, 6.667; 95% CI, 2.279-19.504; P<0.01). Also, within the DM group, the decreased POMA score was associated with multiple diabetic complications (OR, 5.977; 95% CI, 1.378-25.926; P<0.05), decreased IADL score (OR, 10.288; 95% CI, 2.410-43.915; P<0.01), and MMSE score (OR, 13.757; 95% CI, 2.556-74.048; P<0.01). CONCLUSION: Multiple diabetic complications, lower MMSE, ADL, and IADL scores were associated with declining balance function in the older persons with DM. These findings can alert physicians to detect and intervene earlier on declining balance in older persons with DM.

Apolipoprotein E gene polymorphisms and risk for coronary artery disease in Chinese Xinjiang Uygur and Han population.

OBJECTIVE: To examine the relationship between apolipoprotein E (Apo E) gene polymorphism and risk of coronary artery disease (CAD), analyzing association of polymorphism with classical risk factors. METHODS: A total of 124 patients (including 84 Han population and 40 Uygur population) with angiographically verified CAD or myocardial infarction were prospectively evaluated. Data referring to hypertension, diabetes, and tobacco consumption were recorded. The levels of total cholesterol (TC), high density lipoprotein (HDL) cholesterol, Apo A1 and B, and triglycerides (TG) were determined. DNA was obtained from 124 patients and 70 controls. In order to determine Apo E genotypes, DNA was PCR amplified and digested with HhaI. The genetic polymorphism of Apo E is due to three common alleles, epsilon (epsilon) 2, epsilon3, epsilon4, at a single autosomal gene locus. These alleles determine the six phenotypes E2/2, E3/3, E4/4, E4/2, E4/3, and E3/2. RESULTS: In Uygur population, the frequency of the epsilon2, epsilon3, and epsilon4 was 0.155, 0.648, and 0.197 respectively. In Han population, the frequency of the epsilon2, epsilon3, and epsilon4 was 0.081, 0.772, and 0.146 respectively. In the patient group, the frequency of the epsilon2, epsilon3, and epsilon4 was 0.060, 0.758, and 0.182 respectively. In the control group, the frequency of the epsilon2, epsilon3, and epsilon4 was 0.193, 0.671, and 0.136 respectively. epsilon2 frequency of Uygur' patients and controls was 0.050 and 0.290 respectively. Serum low density lipoprotein (LDL) cholesterol, TC, and TG values tended to decrease from the Apo E-4 phenotypes to Apo E-2 phenotypes. When deletion polymorphism of epsilon2 was compared with the common risk factors for CAD, its risk ratio (RR) is 4.38. CONCLUSIONS: These studies confirm and find that Apo E phenotype distribution in Uygur population differs significantly from that in Han population in Xinjiang. CAD patients have significantly lower epsilon2 allele and slightly higher epsilon3 or epsilon4 allele frequency than controls, especially in Uygur population. It shows protective effects of epsilon2 on CAD.