The effect of noninvasive positive pressure ventilation on the expression of ubiquitin-proteasome of skeletal muscle in patients with AECOPD / 中国应用生理学杂志

<p><b>OBJECTIVE</b>To investigate the effect of noninvasive positive pressure ventilation( NIPPy) on the gene and protein expression of biquitin-proteasome of skeletal muscle in patients with acute exacerbation of chronic obstructive pulmonary disease(AECOPD).</p><p><b>METHODS</b>Seven patients with AECOPD by NIPPV were used as the study group, meanwhile, 6 patients with AECOPD who refused NIPPV was the control group. The blood gas analysis, heart rate (HR) and mean arterial pressure (MBp) were monitored before and 14 days after treatment. A skeletal muscle biopsy was performed after 14 days of therapy. The mRNA expression of ribosomal protein S21 (RPS21), Ubiquitin, Ubiquitin combined with enzyme E2 (E2), Ubiquitin ligase E3 (E3) in skeletal muscle cell were measured by RT-PCR. The protein expression of mitochondrial aconitase (AC02), protease C3 (C3), ribosomal protein SLC16 (SLC16) were detected by Western blot.</p><p><b>RESULTS</b>Forteen days after treatment, the patients in NIPPV group got much better improvement in PaCO2, PaO2 and HR than that of the patients.in the control group (P < 0.05). The gene expression of RPS21,Ubiquitin, E2 and E3 in skeletal muscle cell on patients with NIPPV were obviously lower than that of the control group (P < 0.05, P < 0.01). Compared with that of the control group, the protein expression of C3 and AC2 increased significantly in the NIPPV group (P < 0.01). The protein expression of SLC16 was significantly lowered in the treated group (P < 0.01).</p><p><b>CONCLUSION</b>NIPPV can ameliorate the proteasome pathway and energy metabolic disorders in patients with AECOPD.</p>

The effect of bone marrow mesenchymal stem cell transplantation on hypoxic pulmonary hypertension in rats / 中国应用生理学杂志

<p><b>OBJECTIVE</b>To study the influence of bone marrow mesenchymal stem cells (MSCs) transplantation on hypoxic pulmonary hypertension (HPH) in rats.</p><p><b>METHODS</b>SD rats MSCs were separated, cultivated, identified and labeled by the green fluorescence protein (GFP) gene virus and transplanted in vitro. Healthy male SD rats were randomly divided into four groups: Normal control group (NC group) and HPH group (eight rats respectively), HPH+ MSCs transplantation group and HPH+ VEGF+ MSCs transplantation group (twenty-four respectively). The test employed atmospheric intermittent low oxygen method to establish the rat model of pulmonary hypertension and stem cells were transferred and transplanted. The rats' mean pulmonary artery pressure (mPAP) was observed; right ventricular hypertrophy index (RVHI) was calculated; the morphological change of lung small artery in various groups of rats was observed under the microscope; the distribution of lung small artery and adenovirus transfection fluorescently labeled MSCs was observed under a fluorescent microscope after 7, 14 and 28 days when stem cell was transplanted.</p><p><b>RESULTS</b>For NC group, the mPAP (mmHg) was 15.5 +/- 1.5 after twenty-eight days while the mPAPs for HPH , MSCs and MSCs+ VEGF were 26.1 +/- 1.9, 21.6 +/- 2.7 and 20.1 +/- 2.9 respectively which were apparently higher than that of NC group (P < 0.01) and compared with HPH group (P < 0.01), which declined clearly. There was no significant difference between MSCs and MSCs+ VEGF. After twenty-eight days, RVHI for NC group was 0.28 +/- 0.02 while the RVHI for HPH, MSCs and MSCs + VEGF were 0.43 +/- 0.07, 0.34 +/- 0.03 and 0.35 +/- 0.01 respectively which was apparently higher than that of NC group (P < 0.01) but which was clearly lower than that of MSCs and MSCs+ VEGF (P < 0.05) and there was no significant difference between MSCs and MSCs + VEGF. For HPH group, pulmonary arteriole wall became apparently thicker, the lumen became significantly narrow and nearly obstructed after twenty-eight days, the endothelial cells were incomplete; compared with HPH group, pulmonary arteriole wall of MSCs group became thin, the lumen was smooth and the completeness of endothelial cells was improved. Whereas for MSCs and MSCs + VEGF, these changes were not significantly clear.</p><p><b>CONCLUSION</b>After MSCs transplantation, mPAP and RVHI decline sharply and lung small artery remodeling is improved which partially reverses HPH process; there is no significant difference between VEGF together with MSCs transplantation group and pure MSCs.</p>

Serological and molecular capsular typing, antibiotic susceptibility and multilocus sequence typing of Streptococcus pneumoniae isolates from invasive and non-invasive infections / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Streptococcus pneumoniae (S. pneumoniae) is a major causative agent of severe infections, including sepsis, pneumonia, meningitis, and otitis media, and has become a major public health concern. We report the pneumococcal serotype and sequence type (ST) distribution, and antimicrobial resistance of 39 S. pneumoniae strains from seven hospitals in China.</p><p><b>METHODS</b>Blood/cerebrospinal fluid (CSF) and sputum isolates from patients were analyzed to determine S. pneumoniae serotypes by polymerase chain reaction (PCR) and the Neufeld Quellung reaction, the multilocus sequence types (MLST) by PCR and sequencing, and susceptibility to antimicrobial agents by the VITEK Gram Positive Susceptibility Card.</p><p><b>RESULTS</b>A total of 39 isolates were collected including 21 blood/CSF and 18 sputum isolates. Conventional serotyping by the Quellung reaction required 749 reactions. In contrast, PCR based typing needed only 106 PCR reactions. The most frequent serotypes from the blood/CSF isolates were 14 (38.1%), 19A (14.3%), 23F (9.5%), and 18C (9.5%). In the sputum isolates the most frequent serotypes were 19F (33.3%), 23F (16.7%), 19A (11.1%), and 3 (11.1%). The incidence of penicillin resistance in the blood/CSF and sputum isolates was 66.7% and 55.6%, respectively. Statistical analysis showed that patients = 5 years old had a higher resistance to penicillin when they compared with the patients = 65 years old (P = 0.011). Serotypes 14, 19A and 19F were significantly associated with penicillin resistance (P < 0.001). ST320, ST271, and ST876 isolates showed high resistant rates to several antibiotics including penicillin (P = 0.006). All of the isolates of serotype 19A were resistant to both penicillin and erythromycin, and they were all multi-drug resistant (MDR) isolates.</p><p><b>CONCLUSIONS</b>The specificity and sensitivity of multiplex-PCR are good, and this method represents a substantial savings of time and money, and can be widely used in the laboratory and clinical practice. Data from this research showed an extremely high prevalence of penicillin resistance and an increasing prevalence of multi-drug resistant (MDR) rate in S. pneumoniae. A distinctive emergence of serotype 19A was observed which was also associated with the increasing prevalence of antimicrobial resistance. Therefore, nationwide surveillance of pneumococcal resistance and serotypes is strongly warranted.</p>

Preliminary molecular epidemiology of the Staphylococcus aureus in lower respiratory tract infections: a multicenter study in China / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Staphylococcus aureus (S. aureus) remains as an important microbial pathogen resulting in community and nosocomial acquired infections with significant morbidity and mortality. Few reports for S. aureus in lower respiratory tract infections (LRTIs) have been documented. The aim of this study was to explore the molecular epidemiology of S. aureus in LRTIs in China.</p><p><b>METHODS</b>A multicenter study of the molecular epidemiology of S. aureus in LRTIs was conducted in 21 hospitals in Beijing, Shanghai and twelve other provinces from November 2007 to February 2009. All the collected S. aureus strains were classified as minimum inhibitory concentration (MIC), mecA gene, virulence genes Panton-Valentine Leukocidin (PVL) and γ-hemolysin (hlg), staphylococcal cassette chromosome mec (SCCmec) type, agr type, and Multilocus Sequence Typing (MLST).</p><p><b>RESULTS</b>Totally, nine methicillin-sensitive S. aureus (MSSA) and 29 methicillin-resistant S. aureus (MRSA) strains were isolated after culture from a total of 2829 sputums or bronchoalveolar lavages. The majority of MRSA strains (22/29) had a MIC value of ≥ 512 µg/ml for cefoxitin. The mecA gene acting as the conservative gene was carried by all MRSA strains. PVL genes were detected in only one S. aureus strain (2.63%, 1/38). The hlg gene was detected in almost the all S. aureus (100% in MSSA and 96.56% in MRSA strains). About 75.86% of MRSA strains carried SCCmec III. Agr type 1 was predominant (78.95%) among the identified three agr types (agr types 1, 2, and 3). Totally, ten sequence type (ST) of S. aureus strains were detected. A new sequence type (ST1445) was found besides confirming ST239 as the major sequence type (60.53%). A dendrogram generated from our own MLST database showed all the bootstrap values ≤ 50%.</p><p><b>CONCLUSION</b>Our preliminary epidemiology data show SCCmec III, ST239 and agr type 1 of S. aureus as the predominant strains in LRTIs in Mainland of China.</p>

Drug-resistant genes carried by Acinetobacter baumanii isolated from patients with lower respiratory tract infection / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Acinetobacter baumanii (A. baumanii ) remains an important microbial pathogen resulting in nosocomial acquired infections with significant morbidity and mortality. The mechanism by which nosocomial bacteria, like A. baumanii, attain multidrug resistance to antibiotics is of considerable interest. The aim in this study was to investigate the spread status of antibiotic resistance genes, such as multiple β-lactamase genes and aminoglycoside-modifying enzyme genes, from A. baumanii strains isolated from patients with lower respiratory tract infections (LRTIs).</p><p><b>METHODS</b>Two thousand six hundred and ninety-eight sputum or the bronchoalveolar lavage samples from inpatients with LRTIs were collected in 21 hospitals in the mainland of China from November 2007 to February 2009. All samples were routinely inoculated. The isolated bacterial strains and their susceptibility were analyzed via VITEK-2 expert system. Several kinds of antibiotic resistant genes were further differentiated via polymerase chain reaction and sequencing methods.</p><p><b>RESULTS</b>Totally, 39 A. baumanii strains were isolated from 2698 sputum or bronchoalveolar lavage samples. There was not only a high resistant rate of the isolated A. baumanii strains to ampicillin and first- and second-generation cephalosporins (94.87%, 100% and 97.44%, respectively), but also to the third-generation cephalosporins (ceftriaxone at 92.31%, ceftazidine at 51.28%) and imipenem (43.59%) as well. The lowest antibiotic resistance rate of 20.51% was found to amikacin. The OXA-23 gene was identified in 17 strains of A. baumanii, and the AmpC gene in 23 strains. The TEM-1 gene was carried in 15 strains. PER-1 and SHV-2 genes were detected in two different strains. Aminoglycoside-modifying enzyme gene aac-3-Ia was found in 23 strains, and the aac-6'-Ib gene in 19 strains. aac-3-Ia and aac-6'-Ib genes hibernated in three A. baumanii strains that showed no drug-resistant phenotype.</p><p><b>CONCLUSIONS</b>A. baumanii can carry multiple drug-resistant genes at the same time and result in multi-drug resistance. Aminoglycoside-modifying enzyme genes could be hibernating in aminoglycoside sensitive strains without expressing their phenotype.</p>

The expression of alpha-smooth muscle actin in primary cultural fibroblasts of rats / 中国应用生理学杂志

<p><b>AIM</b>To observe the expression of a-smooth muscle actin(a-SMA) in primary cultural fibroblasts of rats.</p><p><b>METHODS</b>12 female Wistar rats were randomly assigned into two groups, the normal group and the model group. The model group was filled with bleomycin A2 (5 mg/kg) once into the trachea. The normal group was filled with equal saline into the trachea. The rats were sacrificed under drugged state at 28 days of feeding, then Hematoxylin-Eosin staining and electron microscopy were used to evaluate the foundation of the model. The isolated fibroblasts from the rats were cultured in vitro. Flow cytometry was used in the test to observe the expression of alpha-SMA in fibroblast in vitro in rats.</p><p><b>RESULTS</b>The formation of fibroblast foci was observed in the model group by optical microscope. The ultrastructure in pulmonary tissue of the model group rats were changed and proliferated myofibroblasts with filaments were found in the alveolar septa by electron microscopy. The expression of alpha-SMA was positive in the normal and model group. There was no difference between the two groups in the rates of positive cells (P > 0.05).</p><p><b>CONCLUSION</b>Both the normal and model groups had the phenotype conversion in lung fibroblasts in vitro.</p>

Monoclonal antibodies against human tumor metastasis suppressor gene-1 TMSG-1: preparation, characterization and application / 中华病理学杂志

<p><b>OBJECTIVE</b>In order to clarify the exact molecular weight of tumor metastasis suppressor gene-1 (TMSG-1) protein and its cellular localization, a monoclonal antibody against TMSG-1 was prepared, characterized and applied to evaluate the metastatic potential of human tumors.</p><p><b>METHODS</b>A dominant epitope-TMSG-1(15)-derived from TMSG-1 was synthesized based on Fmoc method, and the hapten was conjugated to Imject Maleimide activated mcKLH as a carrier protein. The antigen preparation was used to immunize BAL B/C mice. Hybridomas were generated and screened by ELISA for specific monoclonal antibodies, which were further characterized by western blotting and immunohistochemical staining.</p><p><b>RESULTS</b>One hybridoma cell line secreting anti-TMSG-1 antibody, designated as C8, was eventually established after primary ELISA screening, followed by rapid limited dilution procedure. It was confirmed that C8 was of IgM isotype. Result of competitive inhibition assay showed that the antibody was TMSG-1 specific. Using this antibody, an expected protein band of about 45,000 (relative molecular mass) was detected in the non-metastatic variants PC(3)-2B4 and PG-LH7 cells by Western blotting, but not in the isogenetic metastatic variants of PC3-1E8 and PG-BE1 cells. Immunohistochemistry using C8 showed a positive staining of cell membrane and cytoplasm of 2B4 and LH7 cells, whereas 1E8 and BE1 cells were non-reactive. Immunostaining using C8 of paraffin sections of 52 breast carcinomas and 41 colon cancers demonstrated a strong positivity in non-metastatic tumors, but none to weakly reactive in metastatic tumors (P < 0.05).</p><p><b>CONCLUSION</b>C8 monoclonal antibody against the synthetic peptide is TMSG-1 specific and is effective for Western blot and immunohistochemistry assays to detect TMSG-1 expression in cancer cells. TMSG-1 protein is about 45 000 (relative molecular mass) at cell membrane and cytoplasm of tumor cells. Expression of TMSG-1 protein correlates well, inversely with the tumor metastatic potential.</p>

Lung pathology and pathogenesis of severe acute respiratory syndrome: a report of six full autopsies / 中华病理学杂志

<p><b>OBJECTIVE</b>Severe acute respiratory syndrome (SARS) is an emerging infectious disease that first manifested in humans in November 2002. The SARS-associated coronavirus (SARS-CoV) has been identified as the causal agent, but the pathology and pathogenesis are still not quite clear.</p><p><b>METHODS</b>Post-mortem lung samples from six patients who died from SARS from April to July 2003 were studied by light and electron microscopy, Masson trichromal staining and immunohistochemistry. Evidence of infection with the SARS-CoV was determined by reverse-transcription PCR (RT-PCR) , serological examination and electron microscopy.</p><p><b>RESULTS</b>Four of six patients had serological and RT-PCR evidence of recent infection of SARS-CoV. Morphologic changes are summarized as follows: (1) Diffuse and bilateral lung consolidation was seen in all patients (6/6) with increasing lung weight. (2) Diffuse alveolar damage was universal (6/6) with hyaline membrane formation (6/6), intra-alveolar edema/hemorrhage (6/6), fibrin deposition (6/6), pneumocyte desquamation (6/6). A marked disruption in the integrity of the alveolar epithelium was confirmed by immunostaining for the epithelial marker AE1/AE3 (6/6). (3) Type II pneumocytes, with mild hyperplasia, atypia, cytomegaly with granular amphophilic cytoplasm and intracytoplasmic lipid accumulation (5/6). (4) Giant cells in the alveoli were seen in five of 6 patients (5/6) , most of which were positive for the epithelial marker AE1/AE3 (5/6), but some cells were positive for the macrophage marker CD68(2/6). (5) A pronounced increase of macrophages were seen in the alveoli and the interstitium of the lung (6/6), which was confirmed by histological study and immunohistochemistry. (6) Haemophagocytosis was present in five of the 6 patients(5/6). (7) Lung fibrosis was seen in five patients(5/6), with alveolar septa and interstitium thickening(5/6), intraalveolar organizing exudates (6/6) and pleura thickening (4/6). Proliferation of collagen was confirmed by Masson trichromal staining, most of which was type III collagen by immunostaining. The formation of distinctive fibroblast/myofibroblast foci was seen in five patients (5/6) by light microscopy and immunochemistry. (8) Squamous metaplasia of bronchial mucosa was seen in five patients(5/6). (9) Thrombi was seen in all patients(6/6). (10) Accompanying infection was present in two patients, one was bacteria, the other was fungus. In addition, electron microscopy revealed viral particles in the cytoplasm of alveolar epithelial cells and endothelial cells corresponding to coronavirus.</p><p><b>CONCLUSION</b>Direct injury of SARS-CoV on alveolar epithelium, prominent macrophage infiltration and distinctive fibroblast/myofibroblast proliferation may play major roles in the pathogenesis of SARS.</p>