[Determination of tetrahydrofuran in urine by headspace solid-phase microextraction and gas chromatography].

OBJECTIVE: Headspace solid-phase microextraction (HS-SPME) was used pre-concentration procedure for the determination of tetrahydrofuran in urine by gas chromatography with hydrogen flame detector. METHODS: Several parameters controlling SPME was studied and optimised: SPME fiber, extraction time and extraction temperature, desorption time and desorption temperature. RESULTS: Under optimal conditions, the correlation coefficient was 0.9998 and good recoveries (range from 93.0% ∼ 100.8%) were achieved, the detection limit was 0.5 µg/L. CONCLUSION: The method can be applied to the determination of trace amount of tetrahydrofuran in urine.

[Improvement and application of diagnostic techniques of the skeletal muscle biopsy].

OBJECTIVE: To summarize the improvement of various muscle staining techniques and discuss their application in diagnosis of neuromuscular diseases. METHODS: Three hundred cases of skeletal muscle biopsy samples were examined by histopathological methods. The flash-freezing techniques were used for the preparation of frozen section, which were stained with HE, Gomori trichromic (GMR), glycogen (PAS), and fat acid (oil red O); the enzyme histochemical staining with myosin adenosine triphosphatase (ATPase), and NADH-TR. Those stained methods had been improved. The immunohistochemical staining with dystrophin; and transmission electronic microscopy were used. RESULTS: Deepfreeze with heteropentane-liquid nitrogen and flash-freezing techniques could avoid artifacts of ice crystal vacuolation. GMR staining mainly showed the degenerative and necrotic lesions of mitochondria and muscle fibers. Oil red O staining showed the increase of lipid in muscle fibers. PAS staining mainly showed glycogen and glycoprotein. Application of frozen sections in muscular tissue was better than that of paraffin sections. NADH-TR staining and ATPase staining could distinguish the two types of muscle fibers, and show the changes of inner structure of muscle fibers and mitochondria enzymes. CONCLUSION: The distribution and characteristic pathological changes of the two types of muscle fibers can be showed clearly by enzyme and non-enzyme histochemical staining techniques of the skeletal muscle. These methods can compliment with each other. Only after understanding the technical principles can we master and apply these methods.

[Effects of deficit irrigation on water-radiation use and yield of Coffea arabica under different shade cultivation modes in dry-hot region]. / ä¸åŒè«è”½æ ½åŸ¹ä¸‹äºç¼ºçŒæº‰å¯¹å¹²çƒ­åŒºå°ç²’å’–å•¡æ°´å ‰åˆ©ç”¨å’Œäº§é‡çš„å½±å“.

The yield and quality of Coffea arabica cannot be guaranteed due to irrational irrigation and light management in dry-hot region. To investigate the best coupling mode of irrigation and shade cultivation of C. arabica in dry-hot region, a field experiment was carried out with different irrigation levels and shade cultivation modes. Three levels of irrigation, i.e., full irrigation, light deficit irrigation and severe deficit irrigation, and four modes of shade cultivation, i.e., no shade cultivation (C. arabica monoculture), light shade cultivation (intercropping of four-line C. arabica and a line banana), moderate shade cultivation (intercropping of three-line C. arabica and a line banana) and severe shade cultivation (intercropping of two-line C. arabica and a line banana) were designed to examine the effects of deficit irrigation on growth, photosynthetic characteristics of lea-ves, water-radiation use and yield of C. arabica under the shade of banana trees. The results showed that net photosynthesis rate (Pn), transpiration rate (Tr), stomatal conductance (gs), leaf water use efficiency (LWUE), leaf radiation use efficiency (LRUE) increased with the increases of irrigation amount, while the intercellular CO2 concentration (Ci) decreased with the increases of irrigation amount. Compared with full irrigation, light deficit irrigation decreased dry bean yield by 9.4%, severe deficit irrigation decreased dry bean yield and water use efficiency (WUE) by 36.7% and 16.9%, respectively. Pn, Tr, gs, LWUE first increased and then decreased with the increases of shading degree, with moderate shade cultivation having the maximum increment. Compared with no shade cultivation, light shade cultivation increased dry bean yield and WUE by 13.0% and 12.9%, respectively; moderate shade cultivation increased dry bean yield and WUE by 23.1% and 23.4%, respectively. The dry bean yield, WUE, volume and fresh mass of 100-bean of C. arabica increased in different degrees with the increases of irrigation amount and shading degree, with the largest enhancement of dry bean yield and WUE occurring in full irrigation under moderate shade cultivation. Soil water content in the same soil depth decreased with the increases of shading degree. Soil water content in the same place increased first and then decreased with the increases of soil la-yer depth in 0-50 cm soil layer. Results of regression analysis showed that LRUE had a significantly negative exponential relation or Logistic-curve variation with photosynthetically active radiation. Therefore, the suitable combination of irrigation treatment and banana shade cultivation mode for C. arabica was full irrigation and moderate shade cultivation, considering the comprehensive benefit of superior quality and high production and high efficiency water-radiation use.

[Coupling effects of periodic rewatering after drought stress and nitrogen fertilizer on growth and water and nitrogen productivity of Coffea arabica]. / 干旱胁迫-复水与氮肥耦合对小粒咖啡生长和水氮生产力的影响.

The effects of periodic rewatering after drought stress and nitrogen fertilizer on growth, yield, photosynthetic characteristics of leaves and water and nitrogen productivity of Coffea arabica (Katim P7963) were studied under different nitrogen application levels in 2.5 consecutive years. Irrigation (periodic rewatering after drought stress) and nitrogen were designed as two factors, with four modes of irrigation, namely, full irrigation (IF-F: 100%ET0+100%ET0, ET0 was reference crop evapotranspiration), rewatering after light drought stress (IL-F: 80%ET0+100%ET0), rewatering after moderate drought stress (IM-F: 60%ET0+100%ET0) and rewatering after severe drought stress (IS-F: 40%ET0+100%ET0), and three levels of nitrogen, namely, high nitrogen (NH: 750 kg N·hm-2 each time), middle nitrogen (NM: 500 kg N·hm-2 each time), low nitrogen (NL: 250 kg N·hm-2 each time), and nitrogen was equally applied for 4 times. The results showed that irrigation and nitrogen had significant effect on plant height, stem diameter, yield and water and nitrogen productivity of C. arabica, and plant height and stem diameter showed S-curve with the day ordinal number, and leaf photosynthesis decreased significantly under drought stress but most photosynthesis index recovered somewhat after rewatering. Compared with IF-F, IL-F increased dry bean yield by 6.9%, while IM-F and IS-F decreased dry bean yield by 15.2% and 38.5%, respectively; IL-F and IM-F increased water use efficiency by 18.8% and 6.0%, respectively, while IS-F decreased water use efficiency by 12.1%; IL-F increased nitrogen partial productivity by 6.1%, while IM-F and IS-F decreased nitrogen partial productivity by 14.0% and 36.0%, respectively. Compared with NH, NM increased dry bean yield and water use efficiency by 20.9% and 19.3%, while NL decreased dry bean yield and water use efficiency by 42.4% and 41.9%, respectively; NM and NL increased nitrogen partial productivity by 81.4% and 72.9%, respectively. Compared with IF-FNH, IL-FNM increased dry bean yield, water use efficiency and nitrogen partial productivity by 37.6%, 52.9% and 106.4%, respectively. Regression analysis showed that the yield of dry bean was the maximum (2362 kg·hm-2) when the irrigation amount was 318 mm and the nitrogen application amount was 583 kg·hm-2; the water use efficiency was the maximum (0.78 kg·m-3) when the irrigationamount was 295 mm and the nitrogen application amount was 584 kg·hm-2, that's to say when yield of dry bean and water use efficiency reach the maximum value at the same time, the combination was the closest to IL-FNM. Therefore, the best combination of water and nitrogen model for C. arabica was IL-FNM.

[Comparison of three staining methods for hepatitis B virus].

OBJECTIVE: To compare the advantages and disadvantages of three staining methods of HBV. METHODS: Normal Liver tissue and HBV-infected, HCV-infected, or dually infected (HBV and HCV) liver tissues were selected for this study. Formalin-fixed, paraffin-embedded sections (4 microm) were prepared. Each of the liver tissue specimens was detected by three staining methods, including immunohistochemical methods, Shikata's orcein stain and Victoria blue stain,respectively. RESULTS: In the three methods, all of six HBV-infected cases showed intense staining, and three cases with dual infection (HBV and HCV) were weakly positive. However, both normal and HCV-infected liver tissues showed no staining. HBsAg stained dark brown with Immunohistochemical stain; HBsAg containing ground-glass hepatocytes stained magenta with Shikata's orcein stain; HBsAg stained blue with Victoria blue. CONCLUSION: Each of three methods has its own advantages and disadvantages: high specificity and sensitivity, but high cost for immunohistochemical methods; complicated and overelaborate procedure for preparation of solutions, lower specificity and sensitivity, but low cost, for special staining methods.

[Mixed epithelial and stromal tumor of kidney].

OBJECTIVE: To study the clinicopathological features and differential diagnoses of mixed epithelial and stromal tumor of the kidney. METHODS: Clinical and pathological characteristics of 4 cases of mixed epithelial and stromal tumor of the kidney were studied. RESULTS: Three patients were female and one was male. All patients presented with flank pain and hematuria. Radiologic studies revealed cystic and solid masses involving the kidney. Grossly the tumors had a solid and cyst appearance. Microscopically, the tumors were composed of a mixture of stromal and epithelial elements. The epithelial elements were variable in cell types including cuboidal, hobnail and columnar cells. One case showed Müllerian and intestinal epithelial differentiations. Stromal elements essentially consisted of spindle cells, with thick-walled blood vessels and bands of smooth muscle cells as distinctive features of the tumor. Immunohistochemical staining revealed that the epithelial components were positive for AE1/AE3, whereas the stromal components were positive for ER, PR, and SMA. All patients underwent nephrectomy and were well without evidence of recurrence. CONCLUSIONS: Mixed epithelial and stromal tumor of the kidney is a benign neoplasm with distinct histopathological features. It should be distinguished from many other renal neoplasms. Surgical intervention is a preferred therapy.

[Expression of macrophage inflammatory protein-1alpha, a disintegrin-like and metalloproteinase 8 and 12, and CD68 protein in giant cell lesions of jaw and giant cell tumors of long bone].

OBJECTIVE: To detect the expression of macrophage inflammatory protein-1alpha (MIP-1alpha), a disintegrin-like and metalloproteinase (ADAM) 8 and 12 and CD68 protein in giant cell lesions of jaw and giant cell tumors of long bone, and to study their effects on the histogenesis of giant cells in such lesions. METHODS: MIP-1alpha, ADAM8, ADAM12 and CD68 were detected by immunohistochemistry in 24 paraffin-embedded specimens of central giant cell lesions of jaw and giant cell tumors respectively. RESULTS: MIP-1alpha positive signal was located in blood vessels and bone. ADAM8, ADAM12 and CD68 positive signals were located in the cell membrane and cytoplasm of all multinucleated giant cells and some round mononuclear cells in the lesions. In addition, some spindle mononuclear stromal cells were positive for ADAM12 in both lesions. CONCLUSION: Multinucleated giant cells probably originate from CD68-postive round mononuclear cells, which are recruited from monocyte-macrophage system by chemokines, such as MIP-1alpha, followed by cell fusion mediated by ADAM8 and ADAM12.

[Collagen type III glomerulopathy: a morphologic study].

OBJECTIVE: To study the morphologic changes of collagen type III glomerulopathy and to investigate the possible cellular origin for collagen III production. METHODS: Light microscopy, immunofluorescent staining, immunohistochemistry (for collagen I, III and IV and alpha-SMA) and electron microscopy studies on 3 renal biopsy cases of collagen type III glomerulopathy were performed. RESULTS: Two cases presented with nephrotic syndrome, one of which was associated with systemic hypertension. The third case showed renal impairment and renal hypertension. None had any known family history of renal diseases. Light microscopy showed diffuse thickened glomerular basement membrane and expanded mesangium with deposition of weakly PAS-positive homogeneous material not associated with mesangial cell proliferation. Electron microscopy revealed massive collagen fiber deposits in the subendothelial spaces and mesangium. The mesangial cells also contained bundles of microfilaments in the subplasmalemmal regions. Immunohistochemically, the diffuse positivity for type III collagen corresponded to the homogeneous material seen under light microscopy. The staining for type I and IV collagens was negative. Alpha-SMA was expressed in many mesangial cells. CONCLUSIONS: The diagnosis of collagen type III glomerulopathy can be made on the basis of detailed morphologic examination and ancillary investigations. It is possible that activated mesangial cells may be the cellular origin of collagen III.

[Monoclonal antibodies against human tumor metastasis suppressor gene-1 TMSG-1: preparation, characterization and application].

OBJECTIVE: 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. METHODS: 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. RESULTS: 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). CONCLUSION: 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.

[Lung pathology and pathogenesis of severe acute respiratory syndrome: a report of six full autopsies].

OBJECTIVE: 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. METHODS: 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. RESULTS: 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. CONCLUSION: 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.

[Expression of BCL-10 in gastrointestinal mucosa-associated lymphoid tissue lymphoma].

OBJECTIVE: To explore the significance of BCL-10 protein expression in the gastrointestinal mucosa-associated lymphoid tissue (MALT) lymphoma. METHODS: Immunohistochemistry studies were performed using CD20, CD79a, CD3, CD45RO, CD23, CD5, CD10 monoclonal antibodies in 43 cases of gastrointestinal MALT lymphomas, including 25 indolent classical MALT lymphomas and 18 MALT lymphomas with large cell transformation. BCL-10 protein expression was assayed in the tumor cells. RESULTS: In 25 low-grade MALT lymphomas, expression of BCL-10 was found in the nuclei in 10 cases, in both nuclei and cytoplasm 1 case, in cytoplasm 3 cases and no expression 11 cases. In 18 transformed MALT lymphomas, BCL-10 was expressed in the nuclei in 7 cases, in both nuclei and cytoplasm 1 case, in cytoplasm 2 cases, no expression 8 cases. The frequency of BCL-10 expression in nuclei was the highest (44.2%). CONCLUSION: The frequency of BCL-10 expression in nuclei in the gastrointestinal MALT lymphoma is high, indicating that it may associate with the pathogenesis of this entity, and may be helpful to its diagnosis.