Changes in diaphragm contractility in cigarette smoking-exposed and smoking cessation rats are associated with alterations in mitochondrial morphology and homeostasis.

The effects of cigarette smoking (CS) cessation on the diaphragm are unknown, as are the CS-induced diaphragmatic mitochondrial changes. We examined the changes in diaphragm contractility, as well as alterations in mitochondrial morphology, function and homoeostasis during CS exposure and after cessation. Rats were randomly divided into CS exposure and CS cessation groups: 3-month CS (S3), 6-month CS (S6), 6-month CS followed by 3-month cessation (S6N3). The changes in the diaphragm were investigated, including contractile properties, the ultrastructure, mitochondrial function and the expression of markers of mitochondrial homoeostasis. CS caused irreversible histological disruption and functional depression in the lungs, along with significantly declines in diaphragmatic contractility and more severely in extensor digitorum longus muscular contractility. Such declines were recovered after 3-month CS cessation. CS exposure disrupted the diaphragmatic mitochondrial morphology and function (S6), which was significantly alleviated in the S6N3 group. The mitochondrial homoeostasis was depressed (S6), as indicated by the downregulation of Pink1 and Mfn1. Interestingly, the Mfn1 level was recovered after smoking cessation (S6N3). In conclusion, smoking cessation eased CS-induced diaphragmatic dysfunction and mitochondrial deregulation, which are likely associated with deregulated mitochondrial homoeostasis.

The Levels of Oxidized Phospholipids in High-Density Lipoprotein During the Course of Sepsis and Their Prognostic Value.

Purpose: To examine the levels of 1-palmitoyl-2-(5-oxovaleroyl)-sn-glycero phosphatidylcholine (POVPC) and 1-palmitoyl-2-glutaroyl-sn-glycero-phosphatidylcholine (PGPC) (the oxidized phosphatidylcholines) in HDL during the course of sepsis and to evaluate their prognostic value. Materials and Methods: This prospective cohort pilot study enrolled 25 septic patients and 10 healthy subjects from 2020 to 2021. The HDLs were extracted from patient plasmas at day 1, 3 and 7 after sepsis onset and from healthy plasmas (total 81 plasma samples). These HDLs were then subjected to examining POVPC and PGPC by using an ultra-high performance liquid chromatography coupled with tandem mass spectrometry (UHPLC-MS/MS) system. We further measured the levels of 38 plasma cytokines by Luminex and evaluated the correlation of HDL-POVPC level with these cytokines. Patients were further stratified into survivors and non-survivors to analyze the association of HDL-POVPC level with 28-day mortality. Results: Septic patients exhibited significant increase of HDL-POVPC at day 1, 3 and 7 after sepsis onset (POVPC-D1, p=0.0004; POVPC-D3, p=0.033; POVPC-D7, p=0.004, versus controls). HDL-PGPC was detected only in some septic patients (10 of 25) but not in healthy controls. Septic patients showed a significant change of the plasma cytokines profile. The correlation assay showed that IL-15 and IL-18 levels were positively correlated with HDL-POVPC level, while the macrophage-derived chemokine (MDC) level was negatively correlated with HDL-POVPC level. Furthermore, HDL-POVPC level in non-survivors was significantly increased versus survivors at day 1 and 3 (POVPC-D1, p=0.002; POVPC-D3, p=0.003). Area under ROC curves of POVPC-D1 and POVPC-D3 in predicting 28-day mortality were 0.828 and 0.851. POVPC-D1and POVPC-D3 were the independent risk factors for the death of septic patients (p=0.046 and 0.035). Conclusions: HDL-POVPC was persistently increased in the course of sepsis. POVPC-D1 and POVPC-D3 were significantly correlated with 28-mortality and might be valuable to predict poor prognosis.

Reticulocalbin 3 deficiency in alveolar epithelium attenuated LPS-induced ALI via NF-κB signaling.

Acute respiratory distress syndrome (ARDS) is characterized by acute lung injury (ALI) secondary to an excessive alveolar inflammatory response. Reticulocalbin 3 (Rcn3) is an endoplasmic reticulum (ER) lumen protein in the secretory pathway. We previously reported the indispensable role of Rcn3 in type II alveolar epithelial cells (AECIIs) during lung development and the lung injury repair process. In the present study, we further observed a marked induction of Rcn3 in the alveolar epithelium during LPS-induced ALI. In vitro alveolar epithelial (MLE-12) cells consistently exhibited a significant induction of Rcn3 accompanied with NF-κB activation in response to LPS exposure. We examined the role of Rcn3 in the alveolar inflammatory response by using mice with a selective deletion of Rcn3 in alveolar epithelial cells upon doxycycline administration. The Rcn3 deficiency significantly blunted the ALI and alveolar inflammation induced by intratracheal LPS instillation but not that induced by an intraperitoneal LPS injection (secondary insult); the alleviated ALI was accompanied by decreases in NF-κB activation and NLRP3 levels but not in GRP78 and cleaved caspase-3 levels. The studies conducted in MLE-12 cells consistently showed that Rcn3 knockdown blunted the activations of NF-κB signaling and NLRP3-dependent inflammasome upon LPS exposure. Collectively, these findings suggest a novel role for Rcn3 in regulating the alveolar inflammatory response to pulmonary infection via the NF-κB/NLRP3/inflammasome axis and shed additional light on the mechanism of ARDS/ALI.

Different value of HDL-C in predicting outcome of ARDS secondary to bacterial and viral pneumonia: A retrospective observational study.

BACKGROUND: High density lipoprotein-cholesterol (HDL-C) concentration decreases in septic patients and the low level of HDL-C is associated with poor prognosis. However, no study has yet analyzed its prognostic implication specifically in pneumonia-ARDS cohort. OBJECTIVES: To evaluate the prognostic value of HDL-C levels in ARDS patients secondary to bacterial and viral pneumonia. METHODS: This was a retrospective observational study on 108 pneumonia-ARDS patients in RICU from 2017 to 2019. These patients were stratified into bacterial ARDS group (56) and viral ARDS group (52). The primary outcome was the association between HDL-C levels and 28-day mortality. RESULTS: HDL-C levels were statistically lower in bacterial ARDS patients than those in viral ARDS patients (p<0.001). There were statistic negative correlations between HDL-C and APACHE II/SOFA score in bacterial ARDS patients (r=-0.284, p = 0.034 and r=-0.369, p = 0.005), but not in viral ARDS patients (r=-0.103, p = 0.469 and r=-0.225, p = 0.108). ROC analysis demonstrated that HDL-C had superior prediction value for 28-day mortality and identified HDL-C < 0.42 mmol/L was significantly associated with adverse outcomes in bacterial ARDS patients. The low HDL-C was an independent risk factor for death of bacterial ARDS patients (OR 0.027, 95% CI [0.001-0.905], P = 0.044). CONCLUSIONS: HDL-C might be a valuable marker to assess the 28-d mortality for bacterial ARDS patients rather than viral ARDS patients.

Functional, Ultrastructural, and Transcriptomic Changes in Rat Diaphragms with Different Durations of Cigarette Smoke Exposure.

Aims: The aim of the study was to explore the functional and structural changes of the diaphragm and underlying mechanisms in response to 12 or 24 weeks of cigarette smoke (CS) exposure in rats. Materials and Methods: Rats were exposed to CS to develop a COPD model and the rats exposed to room air served as a control group. Rats were randomly divided into four groups: CS12W, CON12W, CS24W, and CON24W. Pulmonary function, lung histopathology, and the contractile properties and ultrastructure of diaphragm muscle were examined in these rats. The changes of transcriptomic profiling of diaphragm muscle were further compared between CS and control rats by the RNA Seq. Results: Both CS groups showed lower FEV0.3/FVC, elevated mean linear intercept (MLI), and reduced mean alveolar numbers (MAN) vs the control groups. The fatigue index (FI) of the diaphragm muscle from the CS12W group, but not CS24W, was significantly increased. Conversely, the force-frequency curves of the diaphragm muscle from the CS24W group, but not CS12W group, were significantly decreased. Consistently, mitochondrial number density (NA) and volume density (Vv) were increased in the CS12W diaphragm muscle, while being decreased in the CS24W group. Furthermore, the diaphragm transcriptomic profiling results showed that genes regulating cell proliferation and energy metabolic activity were un-regulated and genes regulating protein degradation were down-regulated in the CS12W diaphragm, while CS24W diaphragm showed opposite changes. Conclusion: These observations suggested a transition of diaphragm muscle from initial compensatory to decompensatory changes in function, structure, and gene expression during the development of COPD.

The HDL from septic-ARDS patients with composition changes exacerbates pulmonary endothelial dysfunction and acute lung injury induced by cecal ligation and puncture (CLP) in mice.

BACKGROUND: Septic-acute respiratory distress syndrome (ARDS), characterized by the acute lung injury (ALI) secondary to aberrant systemic inflammatory response, has high morbidity and mortality. Despite increased understanding of ALI pathogenesis, the therapies to prevent lung dysfunction underlying systemic inflammatory disorder remain elusive. The high density lipoprotein (HDL) has critical protective effects in sepsis and its dysfunction has a manifested contribution to septic organ failure. However, the adverse changes in HDL composition and function in septic-ARDS patients are large unknown. METHODS: To investigate HDL remodeling in septic-ARDS, we analyzed the changes of HDL composition from 40 patients with septic-ARDS (A-HDL) and 40 matched normal controls (N-HDL). To determine the deleterious functional remodeling of HDL, A-HDL or N-HDL was administrated to C57BL/6 and apoA-I knock-out (KO) mice after cecal ligation and puncture (CLP) procedure. Mouse lung microvascular endothelial cells (MLECs) were further treated by these HDLs to investigate whether the adverse effects of A-HDL were associated with endothelial dysfunction. RESULTS: Septic-ARDS patients showed significant changes of HDL composition, accompanied with significantly decreased HDL-C. We further indicated that A-HDL treatment aggravated CLP induced ALI. Intriguingly, these deleterious effects of A-HDL were associated with pulmonary endothelial dysfunction, rather than the increased plasma lipopolysaccharide (LPS). Further in vitro results demonstrated the direct effects of A-HDL on MLECs, including increased endothelial permeability, enhanced expressions of adhesion proteins and pro-inflammatory cytokines via activating NF-κB signaling and decreased junction protein expression. CONCLUSIONS: Our results depicted the remodeling of HDL composition in sepsis, which predisposes lung to ARDS via inducing ECs dysfunction. These results also demonstrated the importance of circulating HDL in regulating alveolar homeostasis.

Maintenance of spontaneous breathing at an intensity of 60%-80% may effectively prevent mechanical ventilation-induced diaphragmatic dysfunction.

Controlled mechanical ventilation (CMV) can cause diaphragmatic motionlessness to induce diaphragmatic dysfunction. Partial maintenance of spontaneous breathing (SB) can reduce ventilation-induced diaphragmatic dysfunction (VIDD). However, to what extent SB is maintained in CMV can attenuate or even prevent VIDD has been rarely reported. The current study aimed to investigate the relationship between SB intensity and VIDD and to identify what intensity of SB maintained in CMV can effectively avoid VIDD. Adult rats were randomly divided according to different SB intensities: SB (0% pressure controlled ventilation (PCV)), high-intensity SB (20% PCV), medium-intensity SB (40% PCV), medium-low intensity SB (60% PCV), low-intensity SB (80% PCV), and PCV (100% PCV). The animals underwent 24-h controlled mechanical ventilation (CMV). The transdiaphragmatic pressure (Pdi), the maximal Pdi (Pdi max) when phrenic nerves were stimulated, Pdi/Pdi max, and the diaphragmatic tonus under different frequencies of electric stimulations were determined. Calpain and caspase-3 were detected using ELISA and the cross-section areas (CSAs) of different types of muscle fibers were measured. The Pdi showed a significant decrease from 20% PCV and the Pdi max showed a significant decrease from 40% PCV (P<0.05). In vivo and vitro diaphragmatic tonus exhibited a significant decrease from 40% PCV and 20% PCV, respectively (P<0.05). From 20% PCV, the CSAs of types I, IIa, and IIb/x muscle fibers showed significant differences, which reached the lowest levels at 100% PCV. SB intensity is negatively associated with the development of VIDD. Maintenance of SB at an intensity of 60%-80% may effectively prevent the occurrence of VIDD.

High-intensity versus low-intensity noninvasive positive pressure ventilation in patients with acute exacerbation of chronic obstructive pulmonary disease (HAPPEN): study protocol for a multicenter randomized controlled trial.

BACKGROUND: Despite the positive outcomes of the use of noninvasive positive pressure ventilation (NPPV) in patients with acute exacerbation of chronic obstructive pulmonary disease (AECOPD), NPPV fails in approximately 15% of patients with AECOPD, possibly because the inspiratory pressure delivered by conventional low-intensity NPPV is insufficient to improve ventilatory status for these patients. High-intensity NPPV, a novel form that delivers high inspiratory pressure, is believed to more efficiently augment alveolar ventilation than low-intensity NPPV, and it has been shown to improve ventilatory status more than low-intensity NPPV in stable AECOPD patients. Whether the application of high-intensity NPPV has therapeutic advantages over low-intensity NPPV in patients with AECOPD remains to be determined. The high-intensity versus low-intensity NPPV in patients with AECOPD (HAPPEN) study will examine whether high-intensity NPPV is more effective for correcting hypercapnia than low-intensity NPPV, hence reducing the need for intubation and improving survival. METHODS/DESIGN: The HAPPEN study is a multicenter, two-arm, single-blind, prospective, randomized controlled trial. In total, 600 AECOPD patients with low to moderate hypercapnic respiratory failure will be included and randomized to receive high-intensity or low-intensity NPPV, with randomization stratified by study center. The primary endpoint is NPPV failure rate, defined as the need for endotracheal intubation and invasive ventilation. Secondary endpoints include the decrement of arterial carbon dioxide tension from baseline to 2 h after randomization, in-hospital and 28-day mortality, and 90-day survival. Patients will be followed up for 90 days after randomization. DISCUSSION: The HAPPEN study will be the first randomized controlled study to investigate whether high-intensity NPPV better corrects hypercapnia and reduces the need for intubation and mortality in AECOPD patients than low-intensity NPPV. The results will help critical care physicians decide the intensity of NPPV delivery to patients with AECOPD. TRIAL REGISTRATION: ClinicalTrials.gov, NCT02985918 . Registered on 7 December 2016.

Neutrophil/lymphocyte ratio is helpful for predicting weaning failure: a prospective, observational cohort study.

BACKGROUND: To assess the usefulness of the neutrophil/lymphocyte ratio (NLR), a marker of inflammation and/or stress, for predicting weaning failure in patients receiving invasive mechanical ventilation (IMV), compared to levels of leukocytes and C-reactive protein (CRP). METHODS: This observational prospective cohort study was conducted from July 2013 to December 2016 in an intensive care unit in China, enrolling 269 consecutive patients receiving IMV. Patients underwent a spontaneous breathing trial (SBT) if they were ready to wean, and underwent extubation if they passed the SBT. The evaluated markers were measured immediately prior to SBT, and compared between weaning-failure and weaning-success patients. Receiver-operating characteristic (ROC) curve and logistic regression analyses were used to evaluate the ability of these markers to predict weaning failure. RESULTS: In all, 94 (34.9%) patients failed the weaning process (66 failed SBT and 28 presented with post-extubation respiratory distress). NLR was a better predictor of failure (area under the ROC curve, 0.69; 95% CI, 0.62-0.76) than leukocyte levels (0.60, 0.53-0.67) and CRP values (0.58, 0.51-0.65). NLR >11, leukocyte counts >10.5×109/L, and CRP >58 mg/L prior to weaning had the best combination of sensitivity (73%, 64%, and 63%, respectively), specificity (59%, 55%, and 63%), positive predictive value (49%, 43%, and 48%), negative predictive value (81%, 74%, and 76%), and diagnostic accuracy (64%, 58%, and 63%) for predicting weaning failure. However, only NLR >11 (odds ratio, 5.91; 95% CI, 3.08-11.33; P<0.001) was an independent predictor of weaning failure in the adjusted logistic regression model. CONCLUSIONS: NLR may be a useful marker for predicting weaning failure, and weaning at NLR >11 might be considered with caution. Further study with a larger sample size and with weaning outcome as a variable of concern is warranted. Trial registration: ClinicalTrials.gov identifier: NCT02981589.

Associations of Pulmonary Fibrosis with Peripheral Blood Th1/Th2 Cell Imbalance and EBF3 Gene Methylation in Uygur Pigeon Breeder's Lung Patients.

Background/Aims Pigeon breeder's lung (PBL) results from Th1/Th2 cell imbalance. B cells inhibit the immune activity of Th1, and EBF3 is a key B cell factor. This study explored the relationship between EBF3 and Th1/Th2 imbalance in chronic PBL cases complicated with pulmonary fibrosis (PF). Methods Twenty Uygur PBL+PF patients, 20 pigeon breeders without PBL or PF, and 20 healthy individuals without pigeon breeding history constituted the patient I, negative control, and normal control groups, respectively. Peripheral blood specimens and case backgrounds were collected between June 2016 and March 2017. EBF3 gene methylation was analyzed by matrix assisted laser desorption ionization-time of flight mass spectrometry. To compare different mechanisms of PF progression in PBL, samples from 20 Uygur PBL patients without PF (at acute and sub-acute stages) were collected between October 2017 and February 2018, constituting the patient II group. EBF3 mRNA expression was evaluated by real-time polymerase chain reaction. IFN-γ, IL-4 and IL-10 expression and Th1/Th2 imbalance in PBL were evaluated by enzyme-linked immunosorbent assay and flow cytometry. Results CpG-2 and general methylation rates in the patient I group were lower than those in the control groups (P˂0.017). The level of EBF3 mRNA expression in the patient I group was significantly higher than that in any other group. Compared with the control groups, the patient I group showed a significantly higher level of IL-4, whereas the patient II group showed a significantly lower level. IL-10 was also expressed more highly in the patient I group than in any other group (P< 0.01). Flow cytometry showed INF-γ dominance (Th1 cytokine) in PBL at the acute/sub-acute stage and IL-4 dominance (Th2 cytokine) at the chronic stage after PF occurred. The general methylation rate was negatively correlated with the mRNA level, with the latter being positively correlated with the IL-10 level and number of pigeons bred in the past 3 months. IL-4 expression was negatively correlated with INF-γ but positively correlated with PF area and duration of pigeon breeding history. Conclusions After PF occurs in chronic PBL, the inflammation type changes from Th1 dominance to Th2 dominance. During PBL development, IL-10 increases before IL-4 does, which may be associated with EBF3 hypomethylation and the involvement of B lymphocytes.

The correlation between pulmonary fibrosis and methylation of peripheral Smad3 in cases of pigeon breeder's lung in a Chinese Uygur population.

Smad3 is a key protein in the transforming growth factor-beta (TGF-ß)/Smad signaling pathway, which is involved in fibrosis in many organs. We investigated the relationship between Smad3 gene methylation and pulmonary fibrosis in pigeon breeder's lung (PBL). Twenty Uygur PBL patients with pulmonary fibrosis in Kashi between October 2015 and March 2016 were enrolled. Twenty PBL-free pigeon breeders and 20 healthy non-pigeon breeders enrolled during the same period constituted the negative and normal control groups, respectively. Participants' data and peripheral blood samples were collected, and three Smad3 CpG loci were examined. Distributions of CpG_2 and CpG_4 methylation rates did not differ across groups, whereas distributions of CpG_3 methylation rates were significantly different among the three groups. The CpG_3 methylation rate was significantly lower in the patient group than in the negative control group. Smad3 mRNA expression was significantly higher in the patient group than in the negative control group but did not differ between the two control groups. TGF-ßlevels were significantly higher in the patient group than in either control group (both P<0.01). Smad3 gene methylation and Smad3 mRNA expression were negatively correlated, with a correlation coefficient of -0.84. The number of pigeons bred during the preceding three months was positively correlated with Smad3 mRNA expression, with a correlation coefficient of 0.77. Smad3 gene hypomethylation might promote pulmonary fibrosis in Uygur PBL patients via increased Smad3 mRNA expression. Smad3 methylation, Smad3 mRNA expression and TGF-ß level were correlated with the number of pigeons bred by patients.

Abnormal bone mineral density and bone turnover marker expression profiles in patients with primary spontaneous pneumothorax.

BACKGROUND: To examine the bone mineral density (BMD) and the role of bone biomarkers, including bone formation marker procollagen type I aminoterminal propeptide (PINP) and N-terminal midmolecule fragment osteocalcin (N-MID), bone resorption marker b-C-telopeptides of type I collagen (b-CTX) and tartrate-resistant acid phosphatase 5b (TRACP5b) in the pathogenesis of PSP. METHODS: Eighty-three consecutive primary spontaneous pneumothorax (PSP) patients (PSP group) and 87 healthy individuals (control group) were enrolled in this study. General data, including gender, age, height, weight, and body mass index (BMI), were recorded. Dual-energy X-ray absorptiometry, electrochemiluminescence immunoassay (ECLIA), and ELISA were used to evaluate bone mineral density and expression levels of bone metabolism markers, including PINP, b-CTX, TRACP5b, N-MID, and 25-hydroxyvitamin D (25-OH VD). RESULTS: Mean height was significantly greater in the PSP group compared with the control group, whereas weight and BMI were lower. Patients in the PSP group had significantly lower average bone mineral density, which mainly manifested as osteopenia (11/12, 91.7%); however, only one patient (8.3%) developed osteoporosis. Serum overexpression of PINP, b-CTX, TRACP5b, and N-MID were found in PSP patients. Expression of 25-OH VD was low in PSP patients. Bone resorption markers showed positive linear relationships with bone formation markers in all participants; whereas only TRACP5b expression negatively correlated with 25-OH VD. Expression levels of all bone turnover markers negatively correlated with BMI. Regression analysis identified risk factors of PSP as age, height, weight, and TRACP5b and 25-OH VD expression levels; whereas gender and PINP, b-CTX, and N-MID expression levels were not significantly associated with the onset of PSP. CONCLUSIONS: It had lower bone mineral density in PSP patients. Bone formation marker PINP, N-MID and bone resorption marker b-CTX, TRACP5b were upregulated in PSP patients. 25-OH VD expression was relatively low in this population of PSP patients. Age, height, weight, and expression levels of TRACP5b and 25-OH VD may be risk factors for PSP.

Correlation of macrophage inflammatory protein-1α single gene polymorphisms with the susceptibility to pigeon breeder's lung in chinese uygur population.

OBJECTIVE: To investigate the correlation of macrophage inflammatory protein-1α (MIP-1α) gene single nucleotide polymorphisms (SNP) with the susceptibility to pigeon breeder's lung (PBL) in Chinese Uygur population. METHODS: A total of 92 Uygur from Xinjiang, China were enrolled in the study. Among them, there were 32 patients with PBL, 30 negative controls with history of exposure to pigeons and 30 normal controls without pigeons contact. SNP genotyping for 24 SNPs of MIP-1α were performed. RESULTS: Genotype distribution of MIP-1α SNPs rs1049191, rs1049195, rs3210166, rs1130374 and rs5029407 were significantly different among the three groups (P<0.05). CONCLUSION: MIP-1α SNPs rs1049191, rs1049195, rs3210166, rs1130374 and rs5029407 might have correlation with the susceptibility to pigeon breeder's lung in Chinese Uygur population.

Simvastatin promotes alveolar epithelial cell proliferation and attenuates cigarette smoke­induced emphysema in rats.

Current treatments for chronic obstructive pulmonary disease (COPD) cannot reverse the pathological process of the disease, therefore, the development of novel agents and strategies for COPD treatment is required. The aim of the present study was to investigate the potential therapeutic value of simvastatin (SmSt) in cigarette smoke­induced emphysema in rats. A total of 24 male and female Wistar rats were randomly divided into four groups. The levels of vascular endothelial growth factor (VEGF) in the lung tissues and bronchoalveolar lavage (BAL) fluid of each group were measured using an enzyme­linked immunoassay. The mRNA expression of VEGF was assessed using reverse transcription­quantitative polymerase chain reaction. The protein expression levels of VEGF and proliferating cell nuclear antigen (PCNA) were determined using immunohistochemical assays. Histological scoring revealed that simvastatin reduced the total inflammatory scores significantly more in the simvastatin­treated smoke­exposed group, compared with the smoke exposed (Sm) group. Significant differences in the average inter­alveolar septal wall distance and mean alveolar numbers were also observed between the SmSt and Sm groups. The levels of VEGF in the BAL fluid and lung tissue homogenates of the SmSt group were similar to those in the simvostatin­only (St) and control (CtL) groups, and significantly higher compared with those in the Sm group. The expression of VEGF in the alveolar and bronchial epithelial cells of the SmSt group was similar to that in the CtL group, and significantly higher compared with that of the Sm group. The percentage of PCNA­positive alveolar epithelial cells was significantly higher in the SmSt group compared with the Sm and CtL groups. Simvastatin exerted a significant impact on the expression of VEGF and attenuated cigarette smoke­induced emphysema in rats. Therefore simvastatin may have beneficial effects in patients with COPD.

Rapamycin attenuates bleomycin-induced pulmonary fibrosis in rats and the expression of metalloproteinase-9 and tissue inhibitors of metalloproteinase-1 in lung tissue.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is the most common and devastating form of interstitial lung disease (ILD) in the clinic. There is no effective therapy except for lung transplantation. Rapamycin is an immunosuppressive drug with potent antifibrotic activity. The purpose of this study was to examine the effects of rapamycin on bleomycin-induced pulmonary fibrosis in rats and the relation to the expression of metalloproteinase-9 (MMP-9) and tissue inhibitor of metalloproteinase-1 (TIMP-1). METHODS: Sprague-Dawley rats were treated with intratracheal injection of 0.3 ml of bleomycin (5 mg/kg) in sterile 0.9% saline to make the pulmonary fibrosis model. Rapamycin was given at a dose of 0.5 mg/kg per gavage, beginning one day before bleomycin instillation and once daily until animal sacrifice. Ten rats in each group were sacrificed at 3, 7, 14, 28 and 56 days after bleomycin administration. Alveolitis and pulmonary fibrosis were semi-quantitatively assessed after HE staining and Masson staining under an Olympus BX40 microscope with an IDA-2000 Image Analysis System. Type I and III collagen fibers were identified by Picro-sirius-polarization. Hydroxyproline content in lung tissue was quantified by a colorimetric-based spectrophotometric assay, MMP-9 and TIMP-1 were detected by immunohistochemistry and by realtime quantitative reverse transcriptase polymerase chain reaction (RT-PCR). RESULTS: Bleomycin induced alveolitis and pulmonary fibrosis of rats was inhibited by rapamycin. Significant inhibition of alveolitis and hydroxyproline product were demonstrated when daily administration of rapamycin lasted for at least 14 days. The inhibitory efficacy on pulmonary fibrosis was unremarkable until rapamycin treatment lasted for at least 28 days (P < 0.05). It was also demonstrated that rapamycin treatment reduced the expression of MMP-9 and TIMP-1 in lung tissue that was increased by bleomycin. CONCLUSION: These results highlight the significance of rapamycin in alleviating alveolitis and pulmonary fibrosis, which is associated with decreased expression of MMP-9 and TIMP-1.

[Platelet factor 4 and ß-thromboglobulin in blood plasma of patients with acute exacerbation of chronic obstructive pulmonary disease].

OBJECTIVE: To explore the role of platelet factor 4 (PF4) and ß-thromboglobulin (ß-TG) in patients with acute exacerbation of chronic obstructive pulmonary disease (AECOPD). METHODS: A total of 71 AECOPD patients and 50 chronic obstructive pulmonary disease (COPD) patients within a stable stage were admitted into Beijing Chaoyang Hospital from January 2008 to June 2010. And another 40 healthy volunteers were selected as control group. The data of demographics, arterial blood gas analysis and pulmonary function parameters was collected and analyzed. The plasma levels of PF4 and ß-TG were measured by enzyme-linked immunosorbent assay (ELISA). Platelet count was measured by hematology analyzer. Statistical analysis was used for PF4, ß-TG and platelet count. Spearman rank correlation was used for correlation analysis. RESULTS: No differences in age and gender existed among the AECOPD, stable and control groups. The plasma level of PF4 in the AECOPD group (2.28 µg/L) was significantly higher than that of the stable group (2.01 µg/L) and control group (1.57 µg/L) (both P < 0.05). The level of ß-TG in AECOPD was 2.32 µg/L and it was significantly higher than that of the stable group (1.85 µg/L) and control group (1.29 µg/L) (both P < 0.05). The differences in platelet counts were insignificant between the AEC OPD group ((196 ± 67) ×10(9)/L), stable group ((194 ± 50) ×10(9)/L) and control group ((190 ± 48) ×10(9)/L). AECOPD group was divided into moderate, severe and very severe groups by pulmonary function parameters. The levels of PF4 and ß-TG in very severe group were significantly higher than those in moderate and severe groups (P < 0.05). A significant positive correlation was observed between PF4 and ß-TG (r = 0.518, P < 0.01). The levels of PF4 and ß-TG were negatively correlated with FEV1%, FEV1/FVC and PaO2 (all P < 0.05). CONCLUSION: Abnormal platelet activation exists in AECOPD. And the levels of PF4 and ß-TG may reflect the severity of AECOPD and can be used as the markers of estimating prethrombotic state.

Coagulation and fibrinolytic indices during the first trimester of pregnancy in women with polycystic ovary syndrome: a preliminary study.

OBJECTIVE: To evaluate the levels of coagulation and fibrinolytic markers during the first trimester of pregnancy in women with polycystic ovary syndrome (PCOS) and determine the effects of PCOS and obesity on the levels of these hemostatic markers. METHODS: A cross-sectional study was conducted in Beijing, China, on women with PCOS (n = 50), healthy women (n = 50), pregnant women with PCOS (n = 50), and healthy pregnant women (n = 50) at 12 weeks of pregnancy. Coagulation and fibrinolytic parameters were measured. RESULTS: The interaction between PCOS and pregnancy appears to exert effects on the activities of coagulation factors VIII and X. The interaction between PCOS and obesity also seems to affect the level of von Willebrand factor. CONCLUSIONS: Pregnant women with PCOS, especially women who are obese, are observed to be in a more prohemostatic state during the first trimester.

Determination of PKC isoform-specific protein expression in pulmonary arteries of rats with chronic hypoxia-induced pulmonary hypertension.

BACKGROUND: Evidence indicates that protein kinase C (PKC) plays a pivotal role in hypoxia-induced pulmonary hypertension (PH), but PKC isoform-specific protein expression in pulmonary arteries and their involvement in hypoxia-induced PH are unclear. MATERIAL/METHODS: Male SD rats (200-250 g) were exposed to normobaric hypoxia (10% oxygen) for 1, 3, 7, 14 and 21 d (days) to induce PH. PKC isoform-specific membrane translocation and protein expression in pulmonary arteries were determined by using Western blot and immunostaining. RESULTS: We found that only 6 isoforms of conventional PKC (cPKC) α, ßI and ßII, and novel PKC (nPKC) δ, ε and η were detected in pulmonary arteries of rats by Western blot. Hypoxic exposure (1-21 d) could induce rat PH with right ventricle (RV) hypertrophy and vascular remodeling. The cPKCßII membrane translocation at 3-7 d and protein levels of cPKCα at 3-14 d, ßI and ßII at 1-21 d decreased, while the nPKCδ membrane translocation at 3-21 d and protein levels at 3-14 d after hypoxic exposure in pulmonary arteries increased significantly when compared with that of the normoxia control group (p<0.05 vs. 0 d, n=6 per group). In addition, the down-regulation of cPKCα,ßI and ßII, and up-regulation of nPKCδ protein expressions at 14 d after hypoxia were further confirmed by immunostaining. CONCLUSIONS: This study is the first systematic analysis of PKC isoform-specific membrane translocation and protein expression in pulmonary arteries, suggesting that the changes in membrane translocation and protein expression of cPKCα, ßI, ßII and nPKCδ are involved in the development of hypoxia-induced rat PH.

[Effects of aerosolized earthworm fibrinolytic enzyme on bleomycin induced pulmonary fibrosis in rats].

OBJECTIVE: To explore the effects of aerosolized earthworm fibrinolytic enzyme (EFE) on bleomycin-induced pulmonary fibrosis in rats. METHODS: A total of 72 male SD rats were divided randomly into 3 groups of bleomycin (BLM) group with intratracheal BLM (5 mg/kg), control group with the same dose of normal saline, then after both receiving aerosolization of normal saline once daily instead of EFE, EFE group with EFE (2500 U/kg) by aerosolization once daily after BLM instillation. Lung histopathology, immunohistochemistry for transforming growth factor ß(1) (TGF-ß(1)), lung hydroxyproline contents, levels of urokinase PA (uPA), tissue plasminogen activator (tPA) and PA inhibitor 1 (PAI-1) in lung and blood were observed at Days 7, 14 and 28 of experiment, respectively. RESULTS: Compared with BLM group, pulmonary fibrosis improved and the TGF-ß(1) expression of lung tissue decreased (P < 0.01). Hydroxyproline content of lung tissue decreased in EFE group compared with BLM group ((5.8 ± 2.5) vs (9.6 ± 1.3), (6.7 ± 1.4) vs (9.7 ± 1.5), (7.5 ± 1.2) vs (9.7 ± 1.4) mg/L, P < 0.01). Compared with BLM group, the uPA levels of lung were elevated in EFE group at Days 7 and 14 ((1.04 ± 0.36) vs (0.72 ± 0.11), (0.90 ± 0.09) vs (0.75 ± 0.08) µg/L, P < 0.05). Moreover, the plasma levels uPA of increased at Days 14 and 28 ((0.32 ± 0.04) vs (0.25 ± 0.02), (0.36 ± 0.05) vs (0.28 ± 0.04) µg/L, P < 0.05). Consistently, compared with BLM group, the tPA levels of lung increased in EFE group ((4.70 ± 0.87) vs (3.01 ± 0.62), (5.72 ± 0.37) vs (3.00 ± 0.51), (6.73 ± 1.12) vs (3.18 ± 0.38) µg/L, P < 0.01) and the plasma levels of tPA also increased ((3.40 ± 0.36) vs (1.79 ± 0.38), (3.17 ± 0.37) vs (2.18 ± 0.17), (3.85 ± 0.56) vs (2.80 ± 1.06) µg/L, P < 0.01). However, compared with BLM group, the PAI-1 levels of lung decreased in EFE group ((6.04 ± 0.81) vs (8.52 ± 1.01), (6.78 ± 0.81) vs (9.81 ± 1.73), (7.63 + 0.99) vs (11.44 ± 2.54), P < 0.05) and the plasma levels of PAI-1 also decreased in EFE group ((4.82 ± 0.42) vs (6.89 ± 0.84), (5.73 ± 0.40) vs (7.30 ± 1.09), (5.64 ± 0.87) vs (7.98 ± 1.10) µg/L, P < 0.05). CONCLUSIONS: Earthworm fibrinolytic enzyme may decrease bleomycin-induced pulmonary fibrosis and TGF-ß(1) expression while increasing fibrinolytic activation. And fibrinolytic strategies are probably useful for the therapy of fibrotic lung diseases.