A high performance liquid chromatography-tandem mass spectrometry(HPLC-MS/MS) method was established for simultaneous determination of seven characteristic components from the active fraction of Alpiniae Officinarum Rhizoma in rat plasma, including galangin, kaempferol, kaempferide, pinocembrin, 1,7-diphenyl-4-en-3-heptanone, 5-hydroxy-7-(4-hydroxy-3-methoxyphenyl)-1-phenyl-3-heptanone(DHPA), and 7-(4-hydroxy-3-methoxyphenyl)-1-phenyl-4-en-3-heptanone(DPHB). The new developed HPLC-MS/MS method was applied to study the pharmacokinetics of the 7 characteristic components in rats with Helicobacter pylori gastritis. A Waters Sunfire C_(18) column(2.1 mm×150 mm, 3.5 μm) was used. The acetonitrile-aqueous solution(containing 0.1% formic acid) was adopted as the mobile phase for gradient elution. Seven components and internal standard(chlorogenic acid) were separated within 12 min. Mass spectrometric detection was performed in multiple reaction monitoring(MRM) mode using electrospray ionization(ESI) source with fast switching between positive and negative ions. The method was verified by specificity, linearity, precision, accuracy, recovery, matrix effect, and stability and met the requirements of pharmacokinetic study on the 7 components in rat plasma. Pharmacokinetic results showed that the average peak time(T_(max)) of the 7 components was 0.31-2.19 h, their elimination half-life(t_(1/2)) was 5.26-16.65 h, and the average residence time(MRT) was 6.29-31.03 h after the oral administration of the active fraction of Alpiniae Officinarum Rhizoma to rats with H. pylori gastritis. The plasma exposure levels of galangin and DHPA were higher than those of the other components. The concentration-time curves of four detected flavonoids showed obvious double peaks. This study elucidated the pharmacokinetic characteristics of 7 characteristic components from the active fraction of Alpiniae Officinarum Rhizoma in rats with H. pylori gastritis, providing a scientific basis for the identification of the pharmacodynamic substances of Alpiniae Officinarum Rhizoma for treatment of H. pylori gastritis and the clinical application of Alpiniae Officinarum Rhizoma in the prevention and treatment of H. pylori gastritis.
Animals ; Rats ; Chromatography, High Pressure Liquid/methods* ; Tandem Mass Spectrometry/methods* ; Drugs, Chinese Herbal/administration & dosage* ; Male ; Helicobacter pylori/drug effects* ; Alpinia/chemistry* ; Rats, Sprague-Dawley ; Gastritis/metabolism* ; Helicobacter Infections/metabolism* ; Flavonoids/blood* ; Rhizome/chemistry* ; Liquid Chromatography-Mass Spectrometry

ObjectiveTo observe the effect of Youguiwan on the leptin/Janus kinase 2 （JAK2）/signal transducer and activator of transcription 3 （STAT3） signaling pathway in the lung tissue of the rat model of chronic obstructive pulmonary disease （COPD） due to kidney-Yang deficiency. MethodForty rats were modeled for COPD with the syndrome of kidney-Yang deficiency by intratracheal instillation of lipopolysaccharide on day 1 and day 14 and continuous fumigation for 6 weeks， during which hydrocortisone was injected intramuscularly at an interval of 3 days. The modeled rats were randomized into model， high- （11.7 g·kg^-1）， medium- （5.85 g·kg^-1）， and low-dose （2.93 g·kg^-1） Youguiwan， and aminophylline （0.054 g·kg^-1） group. In addition， 8 SD rats were set as the blank group. After the completion of modeling， the rats in each group were administrated with the corresponding drug by gavage for 28 consecutive days. After the last administration， samples were collected. A lung function analyzer was used to evaluate the lung function of rats. Enzyme-linked immunosorbent assay was employed to measure the levels of interleukin-17A （IL-17A）， interleukin-6 （IL-6）， and tumor necrosis factor-α （TNF-α） in the bronchoalveolar lavage fluid （BALF）. Hematoxylin-eosin staining was employed to observe the pathological changes in the lung tissue， and Masson staining was employed to observe the deposition of blue collagen fibers around bronchi in the lung tissue and calculate the inflammation score. The immunofluorescence assay was employed to measure the protein content of collagen type Ⅰ （ColⅠ） and α-smooth muscle actin （α-SMA） in the bronchi. The protein and mRNA levels of leptin， IL-17A， JAK2， and STAT3 in the lung tissue were determined by Western blot and real-time fluorescence quantitative polymerase chain reaction， respectively. ResultCompared with the blank group， the model group showed decreased lung function （P<0.01）， elevated levels of IL-6， IL-17A， and TNF-α in the BALF （P<0.01）， and increased lung inflammation score， deposition of subcutaneous collagen fibers in the airway， and ColⅠ and α-SMA proteins （P<0.01）. Furthermore， the modeling up-regulated the proteins and mRNA levels of leptin， IL-17A， JAK2， and STAT3 in the lung tissue （P<0.01） and enhanced the phosphorylation of JAK2 and STAT3 （P<0.01）. Compared with the model group， high- and medium-dose Youguiwan improved the lung function， decreased the inflammation score， reduced collagen fiber deposition and ColⅠ and α-SMA proteins， lowered the levels of IL-6， IL-17A， and TNF-α in the BALF， down-regulated the mRNA and protein levels of leptin， JAK2， STAT3， and IL-17A， and weakened the phosphorylation of JAK2 and STAT3 （P<0.05， P<0.01）. The aminophylline group had higher IL-17A and TNF-α levels than the high-dose Youguiwan group， lower IL-17A level than the medium and low-dose Youguiwan groups， and lower TNF-α level than the low-dose Youguiwan group. Compared with the aminophylline group， the high- and medium-dose Youguiwan groups showed reduced deposition of collagen fibers and protein levels of ColⅠ and α-SMA around the bronchi in the lung tissue （P<0.05， P<0.01）， decreased inflammation score， and down-regulated protein and mRNA levels of leptin， JAK2， STAT3， and IL-17A in the lung tissue. ConclusionYouguiwan can prevent airway remodeling by inhibiting IL-17A to reduce inflammation and collagen deposition in COPD rats， which may be related to the inhibition of the leptin/JAK2/STAT3 signaling pathway.

Objective To explore whether resveratrol (Res) could effectively suppress infarct size or improve neurological deficits in rat models after acute ischemia or ischemia/reperfusion.Methods One hundred and thirty-five rats were divided randomly into nine groups (n=15):sham-operated group,ischemia groups,including 0.04％ propylene glycol solution treatment group (10-6 g/kg,B1 group),low dose of Res intervention group (10-8 g/kg,C1 group),middle dose of Res intervention group (10-7 g/kg,C2 group) and high dose of Res intervention group (10-6 g/kg,C3 group),and ischemia/reperfusion groups,including 0.04％ propylene glycol solution intervention group (10-6 g/kg,B2 group),low dose of Res intervention group (10-8 g/kg,D1 group),middle dose of Res intervention group (10-7 g/kg,D2 group) and high dose of Res intervention group (10-6 g/kg,D3 group).Rats in the sham-operated group only performed separation of right common carotid artery and internal carotid artery;ischemia was induced by middle cerebral artery occlusion; reperfusion was performed 2 h after ischemia; Res intervention was given for 3 days 2 h after reperfusion.Neurological scale scores of all groups were evaluated,infarct volume was measured by 2,3,5-triphenyltetrazolium chloride (TTC) staining,content of water in brain tissue was measured by wet and dry weight method,and the expressions of glutamate/aspartate transporter (GLAST) and glutamate transport-1 (GLT-1) were observed by immunohistochemical method.Results As compared with B groups,the neurological scale scores in the C2,C3,D2 and D3 group were significantly ameliorated with D3 group having the lowest scores (1.30±0.48),infarct volume and brain water content were statistically reduced with D3 group having the smallest infart volume (16.00％ ±6.20％) and lowest water content (52.30％±8.25％),and the expressions of GLAST and GLT-1 were significantly increased with D3 group having the highest levels (39.98±0.77 and 171.76±7.22,P＜0.05).Conclusions Res shows protective effect on cerebral ischemia/reperfusion injury on rats.The mechanism of neuroprotective effect of Res on focal cerebral ischemia/reperfusion injury may be through increasing the expressions of GLAST and GLT-1 in the brain tissue and inhibiting the expression of glutamate,therefore,reducing infarction size and brain edema.

Objective To explore the correlation of lung function and IFN-γ and IL-4 levels in peripheral blood in pulmonary tuberculosis patients. Methods The IFN-γ and IL-4 levels in peripheral blood of 75 pulmonary tuberculosis patients(severe 25 cases,moderate 25 casea,and mild 25 cases) and 30 healthy volunteers were measured by ELISA. Meanwhile FEV1 ,FEV1% and MMEF% of lung function in 75 pulmonary tuberculosis patients were measured.Results The levels of IFN-γ[(0. 204 ±0. 018) μg/L] and IL-4[(0. 523 ±0. 035) μg/L] in peripheral blood were significantly different both between tuberculosis group and control group and among tuberculosis patients ( severe,moderate,and mild) (t =7. 685,6. 374 ,all P ＜0. 05) ;FEV1 ,FEV1% ,and MMEF% of severe and moderate tuberculosis patients were significantly lower than those of mild tuberculosis patients and normal reference value; In tuberculosis patients, FEV1% and MMEF% were negatively related with IL-4 level in peripheral blood( r = -0. 46, -0. 43, all P ＜ 0. 05 ), also significantly positively related with the IFN-γ level in peripheral blood ( r = 0. 47,0. 45, all P ＜0. 05). Conclusion Pulmonary tuberculosis morbility may be due to cellular levels of patients. The IFN-γ and IL-4 levels in peripheral blood could affect the lung function of pulmonary tuberculosis patients.

Objective To provide applied anatomic data for relevant operations of blood vessels of perisacral promontory(BVPSP). Methods The composition of BVPSP including origin, course, diameter of the middle sacral vessels, the distance between the sacral promontory and the sacral 1 transverse trunk were observed on 37 adult cadavers. Result The BVPSP is composed of the common and internal iliac vessels, the superior segment of the middle sacral vessels and the sacral 1 transverse trunk. Middle sacral artery comes from abdominal aorta. Middle sacral veins are thin walled without valves. The average diameter of middle sacral artery and vein is 1.02 mm and 2.53 mm respectively. The distance between the sacral 1 transverse trunk and the sacral promontory is 5.75 mm. Conclusion The composition of BVPSP, especially middle sacral veins, plentiful vascular anastomosis are the anatomical basis leading to massive hemorrhage in the relevant operations.