Missing microbes in infants and children in the COVID-19 pandemic: a study of 1,126 participants in Beijing, China.

The COVID-19 pandemic has caused many fatalities worldwide and continues to affect the health of the recovered patients in the form of long-COVID. In this study, we compared the gut microbiome of uninfected infants and children before the pandemic began (BEFORE cohort, n=906) to that of after the pandemic (AFTER cohort, n=220) to examine the potential impact of social distancing and life habit changes on infant/children gut microbiome. Based on 16S rRNA sequencing, we found a significant change in microbiome composition after the pandemic, with Bacteroides enterotype increasing to 35.45% from 30.46% before the pandemic. qPCR quantification indicated that the bacterial loads of seven keystone taxa decreased by 91.69%-19.58%. Quantitative microbiome profiling, used to enhance the resolution in detecting microbiome differences, revealed a greater explained variance of pandemic on microbiome compared to gender, as well as a significant decrease in bacterial loads in 15 of the 20 major genera. The random forest age-predictor indicated the gut microbiomes were less mature in the after-pandemic cohort than in the before-pandemic cohort in the children group (3-12 years old) and had features of a significantly younger age (average of 1.86 years). Lastly, body weight and height were significantly lower in the after-pandemic cohort than in the before-pandemic cohort in infants (<1 year of age), which was associated with a decrease in bacterial loads in the fecal microbiome.

Meta-analysis of the effectiveness and safety of robotic-assisted versus laparoscopic transabdominal preperitoneal repair for inguinal hernia.

BACKGROUND: Inguinal hernia is a common global disease. This study aims to investigate the effectiveness and safety of robot-assisted transabdominal preperitoneal repair (RTAPP) and laparoscopic transabdominal preperitoneal repair (LTAPP) for inguinal hernia. METHODS: We conducted a thorough search in Cochrane Library, Embase, and PubMed for relevant clinical studies. After applying inclusion and exclusion criteria, the quality of selected studies was assessed using the Jadad scale for randomized controlled studies and the Newcastle-Ottawa scale for observational studies. Meta-analysis was performed using RevMan 5.3 software. RESULTS: A total of ten studies were included, comprising two randomized controlled studies and eight non-randomized controlled studies. Meta-analysis results revealed no statistically significant differences between the RTAPP group and the LTAPP group regarding hospital stay [MD = 0.21 days, 95% CI (-0.09, 0.51), P = 0.17], incidence of seroma [OR = 0.85, 95% CI(0.45, 1.59), P = 0.61], overall complication rate [OR = 1.22, 95% CI(0.68, 2.18), P = 0.51], readmission rate [OR = 1.31, 95% CI(0.23, 7.47), P = 0.76], and recurrence rate [OR = 0.82, 95% CI(0.22, 3.07), P = 0.77]. However, the RTAPP group had longer operation time compared to the LTAPP group [MD = 14.02 minutes, 95% CI (6.65, 21.39), P = 0.0002], and the cost of the RTAPP procedure was higher than that of the LTAPP procedure [MD = $4.17 thousand, 95% CI (2.59, 5.76), P<0.00001]. CONCLUSION: RTAPP for inguinal hernia is a safe and feasible approach, however, it is associated with increased operation time and treatment costs.

Metatranscriptome of human lung microbial communities in a cohort of mechanically ventilated COVID-19 Omicron patients.

The Omicron variant of the severe acute respiratory syndrome coronavirus 2 (SARS­CoV­2) infected a substantial proportion of Chinese population, and understanding the factors underlying the severity of the disease and fatality is valuable for future prevention and clinical treatment. We recruited 64 patients with invasive ventilation for COVID-19 and performed metatranscriptomic sequencing to profile host transcriptomic profiles, plus viral, bacterial, and fungal content, as well as virulence factors and examined their relationships to 28-day mortality were examined. In addition, the bronchoalveolar lavage fluid (BALF) samples from invasive ventilated hospital/community-acquired pneumonia patients (HAP/CAP) sampled in 2019 were included for comparison. Genomic analysis revealed that all Omicron strains belong to BA.5 and BF.7 sub-lineages, with no difference in 28-day mortality between them. Compared to HAP/CAP cohort, invasive ventilated COVID-19 patients have distinct host transcriptomic and microbial signatures in the lower respiratory tract; and in the COVID-19 non-survivors, we found significantly lower gene expressions in pathways related viral processes and positive regulation of protein localization to plasma membrane, higher abundance of opportunistic pathogens including bacterial Alloprevotella, Caulobacter, Escherichia-Shigella, Ralstonia and fungal Aspergillus sydowii and Penicillium rubens. Correlational analysis further revealed significant associations between host immune responses and microbial compositions, besides synergy within viral, bacterial, and fungal pathogens. Our study presents the relationships of lower respiratory tract microbiome and transcriptome in invasive ventilated COVID-19 patients, providing the basis for future clinical treatment and reduction of fatality.

Efficacy and safety of glue mesh fixation for laparoscopic inguinal hernia: A meta-analysis of randomized controlled trials.

It is still controversial whether glue can be used for mesh fixation in laparoscopic inguinal hernia repair. The aim of this meta-analysis was used to systematically evaluate the effectiveness and safety of glue mesh fixation in laparoscopic tension-free inguinal hernia repair. The PubMed database, EMBASE database and Cochrane Library were searched to collect published randomized controlled trials (RCTs) on laparoscopic tension-free inguinal hernia repair with glue mesh fixation. Sixteen RCTs and 2409 patients with inguinal hernia were included. The meta-analysis showed that compared with the mechanical mesh fixation group(MMFG), the glue mesh fixation group(GMFG) had significantly reduced incidences of chronic pain[relative risk (RR) = 0.40, 95% confidence interval (CI) (0.28,0.57), P < 0.00001], urinary retention[RR = 0.53, 95% CI(0.29,0.97), P = 0.04], haematoma[RR = 0.23, 95% CI(0.09,0.58), P = 0.002] and total complications[RR = 0.28, 95% CI(0.18,0.44), P < 0.00001]; there were no significant differences in pain score on postoperative day 1[MD = -1.33, 95% CI(-2.93,0.26), P = 0.10], operation time[MD = 1.46, 95% CI(-3.97,6.88), P = 0.60] and recurrence rate[RR = 0.72, 95% CI(0.35,1.47), P = 0.37] between the two groups. In conclusion, the application of glue mesh fixation in laparoscopic inguinal hernia repair is safe and reliable with fewer complications. Moreover, it can reduce the incidence of chronic pain without increasing the recurrence rate. However, due to the small number of cases in this analysis and limitations in the quality of the included studies, the findings need to be further verified by multicentre, large-sample and high-quality RCTs in the future.

Identification of Enolase 1 as a Potential Target for Magnaporthe oryzae: Integrated Proteomic and Molecular Dynamics Simulation.

Rice blast is an essential factor affecting rice yield and quality, which is caused by Magnaporthe oryzae (M. oryzae). Isobavachalcone (IBC) is a botanical fungicide derived from the seed extract of the Leguminosae plant Psoralea corylifolia L. and has shown an excellent rice blast control effect in field applications. To explore the potential targets of rice blast control, the analysis of the differentially expressed proteins (DEPs) between the liquid culture medium of mycelium treated by 10 mg/L of IBC for 2 h and the control group indicated that Enolase 1 (ENO1) was the most significantly down-regulated DEP with a fold change value of 0.305. In vitro experiments showed that after treating liquid culture mycelium with 10 mg/L of IBC for 0.5, 1, 2, 4, and 8 h, the enzymatic activity of ENO1 in the IBC experimental groups was 0.97, 0.75, 0.52, 0.44, and 0.39 times as much as in the control groups, respectively. To further explore the molecular interaction and binding mode between IBC and ENO1, the three-dimensional structure of ENO1 was established based on homology modeling. Molecular docking and molecular dynamics simulation showed that IBC had a pi-pi stacking effect with the residue TYR_365, a hydrogen bond interaction with the residue ARG_393, and hydrophobic interactions with non-polar residues ALA_361, LYS_362, and VAL_371 of ENO1. These findings indicated that ENO1 is a potential target of M. oryzae, which would pave the way for screening novel effective fungicides targeting ENO1.

The role of prophylactic antibiotics in elective inguinal tension-free hernia repair: A systematic review and meta-analysis.

Whether to use antibiotics to prevent surgical site infection in elective inguinal tension-free hernia repair has been controversial. To systematically evaluate the effect of prophylactic antibiotic application in elective inguinal tension-free hernia repair, we identified all published randomised controlled trials of the effect of prophylactic antibiotic application on elective inguinal tension-free hernia repair were collected by computer retrieval from the China National Knowledge Infrastructure; VIP Database; Wanfang Database; China Biomedical Literature Database; and PubMed, EMBASE and Cochrane Library databases. Meta-analysis was performed by RevMan 5.3 software. The meta-analysis showed that the total incidence of surgical site infections [P = 0.003] and the incidence of superficial surgical site infections [P = 0.004] in the antibiotic group (AG) were lower than those in the non-antibiotic group (NAG). There was no significant difference in the total incidence of postoperative infections [P = 0.06], deep surgical site infections [P = 0.26] and seroma [P = 0.52] between the AG and the NAG. Based on current evidence, the application of prophylactic antibiotics in elective inguinal tension-free hernia repair can prevent the total incidence of surgical site infections and that of superficial surgical site infections but cannot prevent the total incidence of postoperative infection events, incidence of deep surgical site infections and incidence of seroma.

MetaSVs: A pipeline combining long and short reads for analysis and visualization of structural variants in metagenomes.

Structural variants (SVs, including large-scale insertions, deletions, inversions, and translocations) significantly impact the functions of genes in the microbial genome, and SVs in the microbiome are associated with diverse biological processes and human diseases. With the advancements in sequencing and bioinformatics technologies, increasingly, sequencing data and analysis tools are already being extensively utilized for microbiome SV analyses, leading to a higher demand for more dedicated SV analysis workflows. Moreover, due to the unique detection biases of various sequencing technologies, including short-read sequencing (such as Illumina platforms) and long-read sequencing (e.g., Oxford Nanopore and PacBio), SV discovery based on multiple platforms is necessary to comprehensively identify the wide variety of SVs. Here, we establish an integrated pipeline MetaSVs combining Nanopore long reads and Illumina short reads to analyze SVs in the microbial genomes from gut microbiome and further identify differential SVs that can be reflective of metabolic differences. Our pipeline provides researchers easy access to SVs and relevant metabolites in the microbial genomes without the requirement of specific technical expertise, which is particularly useful to researchers interested in metagenomic SVs but lacking sophisticated bioinformatic knowledge.

Dynamic interaction between the human body and the seat during vertical vibration: effect of inclination of the seat pan and the backrest on seat transmissibilities.

Seat inclinations at the seat pan and backrest may affect the sitting comfort. This study was designed to quantify the effect of inclination of a seat pan (0°, 10°, and 20°) and backrest (0°, 15°, and 30°), either foamed or rigid, on the transmissibilities measured at the seat pan and backrest. Seat transmissibilities were measured with fifteen subjects exposed to vertical random vibration between 1 and 15 Hz at 0.5 ms-2 r.m.s. It was found the resonance frequencies in transmissibilities measured at the seat pan and backrest increased with increasing the backrest inclination but were not affected by the seat pan angle. Increasing the foamed backrest inclination increased the peak transmissibilities. Inclination of the rigid seat pan or the rigid backrest reduced the transmissibilities measured at the backrest or the seat pan, respectively. Transmissibilities were more significantly affected by the backrest inclination than the seat pan inclination. Practitioner summary: Seat inclinations may alter the human-seat dynamic interaction and hence the riding discomfort. This study was designed to quantify the effect of inclined seats, either foamed or rigid, on the transmissibilities. It was found the backrest angle affected the transmissibilities more strongly than the seat pan angle.

Identification of Pyruvate Dehydrogenase E1 as a Potential Target against Magnaporthe oryzae through Experimental and Theoretical Investigation.

Magnaporthe oryzae (M. oryzae) is a typical cause of rice blast in agricultural production. Isobavachalcone (IBC), an active ingredient of Psoralea corylifolia L. extract, is an effective fungicide against rice blast. To determine the mechanism of IBC against M. oryzae, the effect of IBC on the metabolic pathway of M. oryzae was explored by transcriptome profiling. In M. oryzae, the expression of pyruvate dehydrogenase E1 (PDHE1), part of the tricarboxylic acid (TCA cycle), was significantly decreased in response to treatment with IBC, which was verified by qPCR and testing of enzyme activity. To further elucidate the interactions between IBC and PDHE1, the 3D structure model of the PDHE1 from M. oryzae was established based on homology modeling. The model was utilized to analyze the molecular interactions through molecular docking and molecular dynamics simulation, revealing that IBC has π-π stacking interactions with residue TYR139 and undergoes hydrogen bonding with residue ASP217 of PDHE1. Additionally, the nonpolar residues PHE111, MET174, ILE 187, VAL188, and MET250 form strong hydrophobic interactions with IBC. The above results reveal that PDHE1 is a potential target for antifungal agents, which will be of great significance for guiding the design of new fungicides. This research clarified the mechanism of IBC against M. oryzae at the molecular level, which will underpin further studies of the inhibitory mechanism of flavonoids and the discovery of new targets. It also provides theoretical guidance for the field application of IBC.

Effect of the thickness of polyurethane foams at the seat pan and the backrest on fore-and-aft in-line and vertical cross-axis seat transmissibility when sitting with various contact conditions of backrest during fore-and-aft vibration.

Ride comfort in vehicles can be affected by seat properties. While previous studies focused on the vertical whole-body vibration, this study was designed to understand the influence of the foam thickness at the seat pan and at the backrest on seat transmissibilities with fore-and-aft vibration. Twelve subjects sitting with or without a vertical backrest were exposed to random fore-and-aft vibration between 1 and 15 Hz with the magnitude of 0.5 ms-2 r.m.s.. It was found that there was no significant difference in the primary resonance frequencies in the fore-and-aft in-line and vertical cross-axis transmissibilities to the seat pan and to the backrest. The resonance frequency in the fore-and-aft in-line transmissibilities to the seat pan and the backrest decreased with increasing thickness of foam at the seat pan and the backrest. Altering the thickness of foam at the seat pan was more effective than changing that at the backrest.

Remote Ischemic Post-Conditioning may Improve Post-Stroke Cognitive Impairment: A Pilot Single Center Randomized Controlled Trial.

BACKGROUND AND PURPOSE: We aimed to demonstrate the tolerability and feasibility and the effect of remote ischemic post-conditioning on cognitive functioning in patients with post-stroke cognitive impairment. METHODS: This was a single-center, randomized, outcome-blinded, placebo-controlled trial, randomized 1:1 to receive 4 cycles of remote ischemic post-conditioning or a sham procedure for 7 days. The primary outcome measure was tolerability and feasibility of remote ischemic post-conditioning. Secondary outcomes to measure the neurological function with national institute of health stroke scale and the cognitive impairment with Montreal Cognitive Assessment scale and Alzheimer's disease assessment scale-cognitive (at baseline, 90 days, 180 days). RESULTS: 48 patients (24 RIPC and 24 Control) were recruited. remote ischemic post-conditioning was well tolerated with 90 out of 96 cycles completed in full. 4 patients experienced vascular events in the control group: 3 cerebrovascular and 1 cardiovascular event versus only 2 cerebrovascular events in the RIPC group. We showed the similar result in the neurological function with national institute of health stroke scale score with no statistically significant differences between RIPC and control group at baseline (P = 0.796) and 90 days (P = 0.401) and 180 days (P = 0.695). But compare with baseline, it was significantly difference in the control and RIPC group at 90 days (P < 0.05) and 180 days (P < 0.05). The comparison of Montreal Cognitive Assessment scale between two groups both showed that P > 0.05 at baseline which was no statistical difference, but P < 0.05 at 90 days and 180 days which were significant statistical difference. The comparison of Alzheimer's disease assessment scale-cognitive between two groups showed that P > 0.05 at baseline (P = 0.955) and 90 days (P = 0.138) was no statistical difference, but P = 0.005<0.05 at 180 days was significant statistical difference. CONCLUSIONS: The remote ischemic post-conditioning for post-stroke cognitive impairment was well tolerated, safe and feasible. The remote ischemic post-conditioning may improve neurological and cognitive outcomes in patients with post-stroke cognitive impairment. A larger trial is warranted. (Clinical Trial Registration-URL: http://www.clinicaltrials.gov. Unique identifier: ChiCTR1800015231.).

Essential oils and ethanol extract from Camellia nitidissima and evaluation of their biological activity.

Camellia nitidissima, a well-known species of yellow Camellia, has undergone commercial cultivation as a new tea resource recently. Herein, the composition, antioxidant and antimicrobial activities of the essential oil and ethanol extract of C. nitidissima were investigated. The essential oils from the leaves and flowers of C. nitidissima were obtained by hydro-distillation. A total of 56 and 34 constituents accounting for 77.5 and 96.8% of the oils were identified by GC-MS. Linalool (35.8%), phytol (7.9%), cis-geranyl acetone (7.3%) and methyl salicylate (6.8%) were found to be the primary components in the leaf oil, while the flower oil was rich in α-eudesmol (34.3%), γ-eudesmol (31.5%) and linalool (11.1%). The ethanol extract of C. nitidissima leaves contained 281.04 mg gallic acid equivalent/g of total phenols. The antioxidant activities of the two oils and extract were evaluated by DPPH and ABTS radical-scavenging assays. The IC50 values varied from 17.4 (extract) to 720.3 µg/mL (flower oil) for DPPH and from 28.8(extract) to 889.6 µg/mL (flower oil) for ABTS. Both essential oils exhibited moderate antioxidant activities, and the extract possessed strong effects close to ascorbic acid. Additionally, the antimicrobial activities of the oils and extract against Staphylococcus aureus, Bacillus subtilis, Escherichia coli and Pseudomonas aeruginosa were evaluated by agar dilution assay. No considerable bactericidal activities were observed for either essential oil or extract compared with ampicillin and tobramycin standards. The results indicated the extract was more efficient than the two essential oils against S. aureus (MIC = 0.625 mg/mL) and B. subtilis (MIC = 1.25 mg/mL).

Upper Limb Ischemic Postconditioning as Adjunct Therapy in Acute Stroke Patients: A Randomized Pilot.

OBJECTIVE: This study aims to observe the clinical effect of upper limb ischemic postconditioning (LIPostC) as an adjunct to treatment with acute stroke patients, possibly due to increased cerebral perfusion. METHODS: We perform a randomized blinded placebo controlled trial in nonthrombolysis patients with acute ischemic stroke, within 72hours of ictus, divided into the LIPostC group and control group. The LIPostC group is induced by 4 cycles of intermittent repeated limb ischemia: alternating 5 minutes inflation (20mm Hg above systolic blood pressure) and 5 minutes deflation performed manually using a standard upper arm blood pressure cuff in the nonparetic arm. The control group receives a sham procedure (cuff inflation to 30mm Hg). Patients underwent the intervention from the time of enrollment to Day 14. Comparison of National Institutes of Health Stroke Scale (NIHSS) score, cerebral infarction volume, relative Perfusion weighted imaging (PWI) parameters (regional relative cerebral blood flow, regional relative mean transit time; preintervention [day 0], day 14, day 90), modified Rankin Scale (mRS; the preintervention score [day 0], the curative ratio at day 90 [we define 0-1 score as close to recovery or full recovery]). RESULTS: Sixty eligible patients with acute stroke (29 LIPostC and 31 control) are recruited age 65years (SD 12.22), blood pressure 156/74mm Hg (SD 14/10), and NIHSS score 5.98 (SD 3.35), mRS score 2.25 (SD .79). Only 1 in the LIPostC group is intolerant the first cycle to give up. All patients tolerate the sham procedure. Two patients experience recurrent stroke versus none in the LIPostC group. Day 90, compared with the control group, there is a significant decrease the NIHSS score, regional relative mean transit time (P < .05) and increase the curative ratio of mRS, regional relative cerebral blood flow(P < .05) in the LIPostC group, which infarct volume decreased by 31.3% (P < .05). CONCLUSIONS: LIPostC after acute stroke is well tolerated and appears safe and feasible. LIPostC may improve neurological outcome, and protective mechanisms may be increased cerebral blood flow to improve cerebral perfusion. A larger trial is warranted.

Different Natural Selection Pressures on the atpF Gene in Evergreen Sclerophyllous and Deciduous Oak Species: Evidence from Comparative Analysis of the Complete Chloroplast Genome of Quercus aquifolioides with Other Oak Species.

Quercus is an economically important and phylogenetically complex genus in the family Fagaceae. Due to extensive hybridization and introgression, it is considered to be one of the most challenging plant taxa, both taxonomically and phylogenetically. Quercus aquifolioides is an evergreen sclerophyllous oak species that is endemic to, but widely distributed across, the Hengduanshan Biodiversity Hotspot in the Eastern Himalayas. Here, we compared the fully assembled chloroplast (cp) genome of Q. aquifolioides with those of three closely related species. The analysis revealed a cp genome ranging in size from 160,415 to 161,304 bp and with a typical quadripartite structure, composed of two inverted repeats (IRs) separated by a small single copy (SSC) and a large single copy (LSC) region. The genome organization, gene number, gene order, and GC content of these four Quercus cp genomes are similar to those of many angiosperm cp genomes. We also analyzed the Q. aquifolioides repeats and microsatellites. Investigating the effects of selection events on shared protein-coding genes using the Ka/Ks ratio showed that significant positive selection had acted on the atpF gene of Q. aquifolioides compared to two deciduous oak species, and that there had been significant purifying selection on the atpF gene in the chloroplast of evergreen sclerophyllous oak trees. In addition, site-specific selection analysis identified positively selected sites in 12 genes. Phylogenetic analysis based on shared protein-coding genes from 14 species defined Q. aquifolioides as belonging to sect. Heterobalanus and being closely related to Q. rubra and Q. aliena. Our findings provide valuable genetic information for use in accurately identifying species, resolving taxonomy, and reconstructing the phylogeny of the genus Quercus.

The pharmacokinetic and pharmacodynamic properties of site-specific pegylated genetically modified recombinant human interleukin-11 in normal and thrombocytopenic monkeys.

In order to improve the pharmacokinetic and pharmacodynamic properties of recombinant human interleukin-11 mutein (mIL-11) and to reduce the frequency of administration, we examined the feasibility of chemical modification of mIL-11 by methoxy polyethylene glycol succinimidyl carbonate (mPEG-SC). PEG-mIL-11 was prepared by a pH controlled amine specific method. Bioactivity of the protein was determined in a IL-11-dependent in vitro bioassay, its pharmacodynamic and pharmacokinetic properties were investigated by using normal and thrombocytopenic monkey models. N-terminus sequencing and peptide mapping analysis revealed that Lys33 is the PEGylated position for PEG-mIL-11. Bioactivity of PEG-mIL-11 assessed by B9-11 cell proliferation assay was comparable to that of mIL-11. More than 79-fold increase in area-under-the curve (AUC) and 26-fold increase in maximum plasma concentration (Cmax) was observed in pharmacokinetic analysis. Single dose administration of the PEG-mIL-11 induced blood platelets number increase and the effect duration were comparable to that of 7 to 10 consecutive daily administration of mIL-11 to the normal and thrombocytopenic monkey models. PEG-mIL-11 is a promising therapeutic for thrombocytopenia.

Hyaluronan synthase 2 regulates fibroblast senescence in pulmonary fibrosis.

Dysregulated repair of lung injury often results in lung fibrosis characterized by unremitting deposition of matrix components including glycosaminoglycan hyaluronan (HA). HA is mainly produced by hyaluronan synthases (HAS) in mesenchymal cells. We previously demonstrated that over-expression of HAS2 in mesenchymal cells in mice regulates the invasiveness of fibroblasts and promotes severe lung fibrosis. The mechanisms that control the resolution of lung fibrosis are unknown. We propose that a critical step in resolving fibrosis is the induction of senescence in fibrotic fibroblasts and hyaluronan synthase 2 may regulate this process. We found that fibrotic fibroblasts developed the characteristics of replicative senescence in culture and that HAS2 expression was dramatically down-regulated. Furthermore, down-regulation of HAS2 initiated and regulated fibroblast senescence through a p27-CDK2-SKP2 pathway. Deletion of HAS2 in mouse mesenchymal cells increased the cellular senescence of fibroblasts in bleomycin-induced mouse lung fibrosis in vivo. These data suggest that HAS2 may be a critical regulator of the fate of pulmonary fibrosis and we propose a model where over-expression of HAS2 promotes an invasive phenotype resulting in severe fibrosis and down-regulation of HAS2 promotes resolution. Targeting HAS2 to induce fibroblast senescence could be an attractive approach to resolve tissue fibrosis.

Oxygen glucose deprivation post-conditioning protects cortical neurons against oxygen-glucose deprivation injury: role of HSP70 and inhibition of apoptosis.

In the present study, we examined the effect of oxygen glucose deprivation (OGD) post-conditioning (PostC) on neural cell apoptosis in OGD-PostC model and the protective effect on primary cortical neurons against OGD injury in vitro. Four-h OGD was induced by OGD by using a specialized and humidified chamber. To initiate OGD, culture medium was replaced with de-oxygenated and glucose-free extracellular solution-Locke's medium. After OGD treatment for 4 h, cells were then allowed to recover for 6 h or 20 h. Then lactate dehydrogenase (LDH) release assay, Western blotting and flow cytometry were used to detect cell death, protein levels and apoptotic cells, respectively. For the PostC treatment, three cycles of 15-min OGD, followed by 15 min normal cultivation, were applied immediately after injurious 4-h OGD. Cells were then allowed to recover for 6 h or 20 h, and cell death was assessed by LDH release assay. Apoptotic cells were flow cytometrically evaluated after 4-h OGD, followed by re-oxygenation for 20 h (O4/R20). In addition, Western blotting was used to examine the expression of heat-shock protein 70 (HSP70), Bcl-2 and Bax. The ratio of Bcl-2 expression was (0.44±0.08)% and (0.76±0.10)%, and that of Bax expression was (0.51±0.05)% and (0.39±0.04)%, and that of HSP70 was (0.42±0.031)% and (0.72±0.045)% respectively in OGD group and PostC group. After O4/R6, the rate of neuron death in PostC group and OGD groups was (28.96±3.03)% and (37.02±4.47)%, respectively. Therefore, the PostC treatment could up-regulate the expression of HSP70 and Bcl-2, but down-regulate Bax expression. As compared with OGD group, OGD-induced neuron death and apoptosis were significantly decreased in PostC group (P<0.05). These findings suggest that PostC inhibited OGD-induced neuron death. This neuro-protective effect is likely achieved by anti-apoptotic mechanisms and is associated with over-expression of HSP70.

Severe lung fibrosis requires an invasive fibroblast phenotype regulated by hyaluronan and CD44.

Tissue fibrosis is a major cause of morbidity, and idiopathic pulmonary fibrosis (IPF) is a terminal illness characterized by unremitting matrix deposition in the lung. The mechanisms that control progressive fibrosis are unknown. Myofibroblasts accumulate at sites of tissue remodeling and produce extracellular matrix components such as collagen and hyaluronan (HA) that ultimately compromise organ function. We found that targeted overexpression of HAS2 (HA synthase 2) by myofibroblasts produced an aggressive phenotype leading to severe lung fibrosis and death after bleomycin-induced injury. Fibroblasts isolated from transgenic mice overexpressing HAS2 showed a greater capacity to invade matrix. Conditional deletion of HAS2 in mesenchymal cells abrogated the invasive fibroblast phenotype, impeded myofibroblast accumulation, and inhibited the development of lung fibrosis. Both the invasive phenotype and the progressive fibrosis were inhibited in the absence of CD44. Treatment with a blocking antibody to CD44 reduced lung fibrosis in mice in vivo. Finally, fibroblasts isolated from patients with IPF exhibited an invasive phenotype that was also dependent on HAS2 and CD44. Understanding the mechanisms leading to an invasive fibroblast phenotype could lead to novel approaches to the treatment of disorders characterized by severe tissue fibrosis.

Airway fibroblasts in asthma manifest an invasive phenotype.

RATIONALE: Invasive cell phenotypes have been demonstrated in malignant transformation, but not in other diseases, such as asthma. Cellular invasiveness is thought to be mediated by transforming growth factor (TGF)-ß1 and matrix metalloproteinases (MMPs). IL-13 is a key T(H)2 cytokine that directs many features of airway remodeling through TGF-ß1 and MMPs. OBJECTIVES: We hypothesized that, in human asthma, IL-13 stimulates increased airway fibroblast invasiveness via TGF-ß1 and MMPs in asthma compared with normal controls. METHODS: Fibroblasts were cultured from endobronchial biopsies in 20 subjects with mild asthma (FEV(1): 90 ± 3.6% pred) and 17 normal control subjects (FEV(1): 102 ± 2.9% pred) who underwent bronchoscopy. Airway fibroblast invasiveness was investigated using Matrigel chambers. IL-13 or IL-13 with TGF-ß1 neutralizing antibody or pan-MMP inhibitor (GM6001) was added to the lower chamber as a chemoattractant. Flow cytometry and immunohistochemistry were performed in a subset of subjects to evaluate IL-13 receptor levels. MEASUREMENTS AND MAIN RESULTS: IL-13 significantly stimulated invasion in asthmatic airway fibroblasts, compared with normal control subjects. Inhibitors of both TGF-ß1 and MMPs blocked IL-13-induced invasion in asthma, but had no effect in normal control subjects. At baseline, in airway tissue, IL-13 receptors were expressed in significantly higher levels in asthma, compared with normal control subjects. In airway fibroblasts, baseline IL-13Rα2 was reduced in asthma compared with normal control subjects. CONCLUSIONS: IL-13 potentiates airway fibroblast invasion through a mechanism involving TGF-ß1 and MMPs. IL-13 receptor subunits are differentially expressed in asthma. These effects may result in IL-13-directed airway remodeling in asthma.

ß-arrestin deficiency protects against pulmonary fibrosis in mice and prevents fibroblast invasion of extracellular matrix.

Idiopathic pulmonary fibrosis is a progressive disease that causes unremitting extracellular matrix deposition with resulting distortion of pulmonary architecture and impaired gas exchange. ß-Arrestins regulate G protein (heterotrimeric guanine nucleotide-binding protein)-coupled receptors through receptor desensitization while also acting as signaling scaffolds to facilitate numerous effector pathways. Here, we examine the role of ß-arrestin1 and ß-arrestin2 in the pathobiology of pulmonary fibrosis. In the bleomycin-induced mouse lung fibrosis model, loss of either ß-arrestin1 or ß-arrestin2 resulted in protection from mortality, inhibition of matrix deposition, and protected lung function. Fibrosis was prevented despite preserved recruitment of inflammatory cells and fibroblast chemotaxis. However, isolated lung fibroblasts from bleomycin-treated ß-arrestin-null mice failed to invade extracellular matrix and displayed altered expression of genes involved in matrix production and degradation. Furthermore, knockdown of ß-arrestin2 in fibroblasts from patients with idiopathic pulmonary fibrosis attenuated the invasive phenotype. These data implicate ß-arrestins as mediators of fibroblast invasion and the development of pulmonary fibrosis, and as a potential target for therapeutic intervention in patients with idiopathic pulmonary fibrosis.