[Electroacupuncture Intervention Improved Pulmonary Function via Promoting Immunoregulation in Chronic Obstructive Pulmonary Disease Rats].

OBJECTIVE: To observe the immunoregulatory effect of electroacupuncture (EA) intervention for muscular dystrophy chronic obstructive pulmonary disease (COPD) rats, so as to investigate its underlying mechanism in improving respiratory function. METHODS: Forty male SD rats were randomly divided into 5 groups: normal, model, EA, exercise, and EA+ exercise (n=8 in each). The muscular dystrophy COPD model was established by placing the rats in a closed box to be exposed to cigarette smoke (3-10 cigarettes/time) for 60 min, twice daily, 6 days a week for 90 days. The EA, exercise and EA+exercise interventions were given beginning from day 80 after exposure to cigarette smoke. EA (2 Hz/40 Hz, 6 mA) was applied to "Danzhong" (CV 17), "Qihai" (CV 6), "Zhongwan" (CV 12), "Liangmen" (ST 21) and bilateral "Quchi" (LI 11) for 10 min, once every other day, for 20 times. The swimming exercise was conducted by forcing the rat to swim in a water box for 10 min, once every other day, for 20 times. The rat's lung function including the resistance of inspiration (RI), functional residual capacity(FRC), pulmonary dynamic compliance (Cdyn), etc., was detected under anesthesia. Pathological changes of the lung tissue were detected by H.E. staining, and the contents of serum TNF-alpha, IL-6 and IL-1 beta assayed by ELISA. RESULTS: After 80 days' exposure to the cigarette smoke, the rats' body weight values in the model, EA, exercise and EA+exercise groups were significantly lower than that of the normal group(P<0.05). Moreover, the RI and FRC levels were significantly increased, and the Cdyn level was remarkably decreased in the model group relevant to the normal group (P<0.01). Following the intervention, both RI and FRC levels were significantly down-regulated in the EA, exercise and EA+exercise groups relevant to the model group (P<0.05), suggesting an improvement of the lung function. But the decreased Cdyn level had no marked improvement in the 3 treatment groups relevant to the model group (P>0.05). The numbers of monocytes and lymphocytes of the lung tissue, and the contents of serum TNF-α, IL-6 and IL-1 ß were significantly higher in the model group than in the normal group (P< 0.01), and significantly lower in the EA, exercise and EA+exercise groups than in the model group (P<0.05), except monocytes in the exercise group (P>0.05). No significant differences were found among the EA, exercise and EA+exercise groups in the levels of RI and FRC, pulmonary monocytes and serum IL-6 and IL-1 ß (P>0.05). The body weight was significantly higher in the exercise and EA+exercise groups than in the EA group, and the pulmonary lymphocytes and serum TNF-α obviously lowered in the EA group than in the exercise group (P<0.05). H.E. staining showed deformation of the bronchial tube cavity, detachment and flattening of the bronchial mucosal epithelial cilia, hyperplasia of Goblet cells, infiltration of abundant inflammatory cells in the submucosal layer and muscular layer, more secretions in the bronchovascular cavity, incomplete alveolar structure, thinning and rupture of the alveolar wall, and expansion of the alveolar cavity to form large pulmonary vesicles after modeling, which was obviously milder in the 3 treatment groups. CONCLUSION: EA intervention can improve the pulmonary function and pathological changes in pulmonary muscular dystrophy COPD rats, which is associated with its effects in reducing pulmonary monocytes and lymphocytes and serum TNF-α, IL-6 and IL-1 ß contents, suggesting an enhancement of immunoregulation.

A facile analytical method for the identification of protease gene profiles from Bacillus thuringiensis strains.

Five pairs of degenerate universal primers have been designed to identify the general protease gene profiles from some distinct Bacillus thuringiensis strains. Based on the PCR amplification patterns and DNA sequences of the cloned fragments, it was noted that the protease gene profiles of the three distinct strains of B. thuringiensis subsp. kurstaki HD73, tenebrionis and israelensis T14001 are varied. Seven protease genes, neutral protease B (nprB), intracellular serine protease A (ispA), extracellular serine protease (vpr), envelope-associated protease (prtH), neutral protease F (nprF), thermostable alkaline serine protease and alkaline serine protease (aprS), with known functions were identified from three distinct B. thuringiensis strains. In addition, five DNA sequences with unknown functions were also identified by this facile analytical method. However, based on the alignment of the derived protein sequences with the protein domain database, it suggested that at least one of these unknown genes, yunA, might be highly protease-related. Thus, the proposed PCR-mediated amplification design could be a facile method for identifying the protease gene profiles as well as for detecting novel protease genes of the B. thuringiensis strains.

The IspA protease's involvement in the regulation of the sporulation process of Bacillus thuringiensis is revealed by proteomic analysis.

We have observed that the process of sporulation of the ispA-deficient mutant was delayed under phase-contrast microscopy. The protein profiles of the ispA-deficient mutant have been analyzed using two-dimensional gel electrophoresis. The results of a proteomic analysis using MALDI-TOF MS indicated that a sporulation-associated protein, pro- [Formula: see text], was upregulated, while two other sporulation-associated proteins, SpoVD and SpoVR, were downregulated in the ispA-deficient mutant. It has been known that pro- [Formula: see text] is a precursor of [Formula: see text] and is required for gene expression related to the late stage of sporulation. Moreover, SpoVD and SpoVR are known to be involved in the formation of the spore cortex. Based on these observations, we propose that the delay in the sporulation process observed in the ispA-deficient mutant may be due to a failure of [Formula: see text] to signal sporulation. This phenomenon may be further enhanced by insufficient amount of SpoVD and SpoVR for cortex formation. In this study, we have revealed for the first time a possible pathway for the regulation of sporulation-associated proteins via IspA.

Dissection of cry gene profiles of Bacillus thuringiensis isolates in Taiwan.

On the basis of the newly revised nomenclature system of cry genes, the PCR amplification method has been adopted to resolve the cry gene combinations of 294 Bacillus thuringiensis isolates from five selected areas of Taiwan. Our results indicate that cry1 (especially cry1A + 1B + 1F) and cry2 were the most abundant cry genes in Taiwan. In contrast, cry3 and cry6 genes were detected only on Yang Ming Mountain, while the cry13 gene was found only on Snow Mountain. In addition, some distinctive combinations of cry genes were detected in distinct areas of Taiwan, such as cry1C, cry1D, cry1C + 1D, cry4, cry1 + 4, cry1 + 11, cry4 + 11, and cry1 + 4 + 11 in the Taipei area; cry1A + 1C + 1F in the Taichung area; cry1E and cry1A + 1B + 1I on Yang Ming Mountain; cry1 + 13, cry1 + 2 + 11, and cry1 + 2 + 13 on Snow Mountain; and cry1 + 5 and cry1 + 2 + 5 on Jade Mountain. These data clearly indicate that the distribution of cry gene combinations of B. thuringiensis isolates seems to be geographically related.