Identification of biomarkers for hepatocellular carcinoma by semiquantitative immunocytochemistry

AIM: To investigate the expression of key biomarkers in hepatoma cell lines, tumor cells from patients’ blood samples, and tumor tissues.

Identification of FXPRLamide Family Neuropeptides from the Japanese Oak Silkworm,Antheraea yamamai Using Immunocytochemistry Methods

In the present study, zooblooting, ELISA, and whole-mount immunocytochemistry methods were used to identify the FXPRLamide family neuropeptides from the Japanese oak silkworm, Antheraea yamamai. The results showed that the genomic DNA from A. yamamai showed positive bands after being hybridized with the fragment of DH-PBAN cDNA from Samia cynthia ricini, which was labeled with [α-32p]-dCTP. The SG showed highest FXPRLamide peptides titer in neural organs. Using an antiserum against Helicoverpa armigera PBAN, PBAN-like immunoreactivity was detected in the SG and TG of A. yamamai by whole-mount immunocytochemistry, and there were three cluster cells in the SG which shows positive PBAN-like immunoreactivity. The titers of FXPRLamide peptides immunoreactivity in the hemolymph were kept at a steady level. During pupation, the titer was increased promptly, but then decreased to a low level after the early pupal stage. The above-mentioned results demonstrate the existence of FXPRLamide family peptides in A. yamamai, but its function needs to be further investigated in the future.

Elucidating the role of SlXTH5 in tomato fruit softening

Fruit softening in tomato(Solanum lycopersicum)is closely associated with cell wall disassembly,which is brought about through the action of a range of cell wall structure-related enzymes and other proteins such as expansins.Xyloglucan endotransglucosylase/hydrolase(XTH)(EC 2.4.1.207 and/or EC 3.2.1.151)has been proposed to be key player involved in xyloglucan metabolism.SlXTH5 showed the highest expression level among all SlXTHs during tomato ripening.In this study,the role of SlXTH5 involved in tomato softening was investigated in CRISPR-based knockout mutants of SlXTH5.Loss-of-function of SlXTH5 in transgenic tomato lines resulted in slightly firmer fruit pericarp,but significantly decreased their color index compared with azygous wild type(WT)control fruits.Increased paste viscosity was detected in CRISPR mutants,indicating that the activity of SlXTH5 is responsible for maintaining cell wall structural integrity.Immunocytochemistry studies were performed using the monoclonal antibody probe LM25 to examine the localization and distribution of xyloglucan in the pericarp cells of the CRISPR mutant fruits.The data indicated more xyloglucan was retained in the pericarp of CRISPR mutant fruit than in WT control fruit.This study revealed the link between SlXTH5 and xyloglucan metabolism and indicated the potential of manipulating SlXTH5 to regulate fruit softening.

Isolation, Cuture and Biological Characteristic Analysis of Stromal and Glandular Epithelial Cells of Buffalo (Bubalus bubalis) Endometrium

[Objective] The experiment aimed to set up a method for isolating and culturing endometrial stromal cells (BESC) and endometrial glandular epithelial cells(BEGEC) of buffalo as well as laid foundation for studying biological mechanism of embryo implantation and uterine diseases. [Method] The enzymatic digestion method, scraping method, serial filtration and differential velocity adherent technique were used to isolate BESC and BEGEC, then immunocytochemical method and TRYPAN-Blue assay were used to determine the purity and survival rate of isolated cells. [Result] The BESC and BEGEC were successfully isolated and cultured while immunocytochemical method and cell count method demonstrated that the purity was over 90%. The result of TRYPAN-Blue assay shown that survival rate of BESC and BEGEC was 91% and 78% respectively. [Conclusion] The enzymatic digestion method, scraping method, serial filtration and differential velocity adherent technique could isolate BESC and BEGEC with high purity.

Localization of Neuropeptidelike Substances During Embryonic Development in Amphibian

The localization of four neuropeptidelike substances during embryonic development in amphibian was studied by using immuno cytochemical technique. The cells with positive reaction appeared firstly in the endoderm cells during early tailbud stage, and then were detected in connective tissue at the outer portion of gastrointestinal tract during tadpole stage. In the nervous system, the cells with positive reaction were observed in cranial ganglion and glial cells at the outer margin of the brain and in the inner wall of ventricles. They were also frequently observed in the epidermis during late tailbud stage. The relationship between the appearance of neuropeptides in timespatial sequences and the development of nervous system, the neural crest origin of the cells with positive reaction, and the role of epidermal conductivity in neuropeptidelike cells in epiderms were discussed.

Optimization of Parameters of Exogenous Gene Mediated by Liposome to Transfect Yak Mammary Epithelial Cells in Vitro

[Objective] The aim of this study was to optimize conditions of exogenous gene mediated by liposome to transfect yak mammary epithelial cells in Vitro.[Method] Yak mammary epithelial cells were isolated and cultivated in Vitro by the methods of collagenase digestion and tissue adhesion.The expression of cytokeratin in yak mammary epithelial cell was detected by immunocytochemistry technique.With green fluorescence protein as the report gene,yak mammary epithelial cells were transfected with exogenous gene m...

Immunocytochemical identification and localization of APUD cells in the gut of seven stomachless teleost fishes

AIM To study the cell types,localization,distribution density and morphology of APUDcells in the intestinal mucosa of stomachlessteleost fishes.METHOD By using the peroxidase-antiperoxidase complex(PAP)immunocytochemical staining technique theidentification,localization and morphology ofimmunoreactive(IR)endocrine cells seattered inthe intestinal mucosa of grass carp(Cyenopharyngodon idellus),black carp(Mylopharyngodon piceus)and common carp(Cyprinus carpio)were investigated with 20kinds of antisera prepared against mammalianpeptide hormones of APUD cells,and likewise byusing avidin-biotin-peroxidase complex(ABC)method those of silver carp(Hypophthalmichthys molitrix),bighead(Aristichthys nobilis),silver crucian carp(Carassius gibelio)and bluntnose black bream(Megalobrama amblyocephala)were alsostudied with 5 different antisera.Thereplacement of the first antiserum by phosphatebuffered saline(PBS)was employed as a control.IR endocrine cells were counted with asquare-mesh ocular micrometer from 10 fieldsselected randomly in every section of each partof the intestine specimen.The average numberof IR endocrine cells per mm2 was counted toquantify their distribution density.RESULT Gastrin(GAS)-,Gastric inhibitorypeptide(GIP)-,glucagon(GLU)-,glucagon-likeimmunoreactants(GLI)-,bovine pancreaticpolypeptide(BPP)-,leucine-enkephalin(ENK)-and substance P(SP)-IR endocrine cells werefound in the gut of grass carp,black carp andcommon carp,and somatostatin(SOM)-IRendocrine cells were only seen in common carp.GAS-,GIP-and GLU-IR endocrine cells werefound in the intestinal mucosa of silver carp,bighead,silver crucian carp and bluntnose blackbream.Most of IR endocrine cells had the higherdistribution density in the foregut and midgut,and were longer in shape.They had a long apicalcytoplasmic process extended to the gut lumenand a basal process extended to adjacent cellsor basement membrane and touched with it.Sometimes,the basal cytoplasmic processformed an enlarged synapse-like structure in thecontiguous part with basement membrane.Thisphenomenon provided new morphologicalevidence for neuroendocrine and paracrinesecretory function of these enteroendocrinecells.CONCLUTION At least 8 kinds of IR endocrinecells were found in the gut of stomachlessteleost species for the first time in China.TheseIR endocrine cells scattering in the gut mucosabelong to the APUD system.Among them,thehormones secreted by SP-,ENK-,SOM-and GLU-IR endocrine cells belong to the peptides of dualdistribution in the brain and gut.This providednew evidence for the concept of brain-gutpeptide.According to the cell types,distribution density,morphologicalcharacteristics and variety in shape of APUDcells in the gut of stomachless teleost fishes,itis deemed that the digestive tract of fishes isalso an endocrine organ of great importance andcomplexity.

Identification,localization and morphology of APUD cells in gastroenteropancreatic system of stomach-containing teleosts

AIM To identify the type localization andmorphology of APUD endocrine cells in thegastroenteropancreatic(GEP)system ofstomach-containing teleosts,and study APUDendocrine system in the stomach,intestine andpancreas of fish species.METHODS Two kinds of immunocytochemical(ICC)techniques of the streptavidin biotin-peroxidase complex(SABC)and streptavidin-peroxidase(S-P)method were used.Theidentification,localization and morphology ofAPUD endocrine cells scattered in the mucosa ofdigestive tract,intermuscular nerve plexus andglandular body of northern snakehead(Channaargus),ricefield eel(Monopterus albus),yellow catfish(Pelteobagrus fulvidraco),mandarinfish(Siniperca chuatsi),largemouthbass(Micropterus salmoides),orientalsheatfish(Silurus asotus),freshwater pomfret(Colossoma brachypomum)and nile tilapia(Tilapia nilotica)were investigated with 8 kindsof antisera.RESULTS The positive reaction of 5-hydroxytryptamine(5-HT)immunoreactiveendocrine(IRE)cells was found in the digestive tract and glandular body of 8 fish species indifferent degree.Only a few gastrin(GAS)-IREcells were seen in C.argus,M.albus and P.fulvidraco.Glucagon(GLU)-IRE cells were notfound in the digestive tract and glandular bodybut existed in pancreatic island of most fishspecies.The positive reaction of growthhormone(GH)-IRE cells was found only inpancreatic island of S.Chuatsi and S.Asotus,no positive reaction in the other 6 fish species.Somatostatin(SOM)-,calcitonin(CAL)-,neurofilament(NF)-and insulin(INS)-IRE cellsin the stomach,intestine and pancreas of 8 kindsof fish were different in distribution and types.The distribution of all 8 APUD cells was the mostin gastrointestinal epithelium mucosa and then indigestive glands.The positive reaction of SOM-and 5-HT-IRE cells was found in intermuscularnerve plexus of intestine of P.fulvidraco andS.chuatsi.Only GH-IRE cells were denselyscattered in the pancreatic islands of S.chuatsiand S.asotus,and odd distribution in thepancreas of S.asotus,SOM-IRE cells weredistributed in the pancreatic islands of S.asotus,C.Brachypomum and T.nilotica.There were INS-IRE cells in the pancreaticislands of S.chuatsi and S.asolus.Eightkinds of APUD cells had longer cell body andcytoplasmic process when they were located inthe gastrointestinal epithelium,and had shortercell body and cytoplasmic process in the gastricgland,and irregular shape in the esophagus andpancreatic island.CONCLUSION Eight kinds of IRE cells were identified in the GEP system of stomach-containing teleosts. These endocrine cells were scattered in gastrointestinal mucosa, intermuscular nerve plexus, gland body, pancreatic gland and islands under APUD system. CAL- and GH-IRE cells in the pancreatic islands of fishes showed functional diversity for these two hormones. Their morphological feature provides evidence of endocrine-paracrine and endocrine-exocrine acting mode. This research can morphologically prove that the GEP endocrine system of fish ( the lowest vertebrate) is almost the same as of mammal and human.

Suppression of P-gp induced multiple drug resistance in a drug resistant gastric cancer cell line by overexpression of Fas

AIM To observe the drug sensitizing effect andrelated mechanisms of fas gene transduction onhuman drug-resistant gastric cancer cellSGC7901/VCR(resistant to Vincristine).METHODS The cell cycle alteration wasobserved by FACS.The sensitivity of gastriccancer cells to apoptosis was determined by invitro apoptosis assay.The drug sensitization ofcells to several anti-tumor drugs was observedby MTT assay.Immunochemical method wasused to show expression of P-gp and Topo Ⅱ ingastric cancer cells.RESULTS Comparing to SGC7901 and pBK-SGC7901/VCR,fas-SGC7901/VCR showeddecreasing G2 cells and increasing S cells,theG2 phase fraction of pBK-SGC7901/VCR wasabout 3.0 times that of fas-SGC7901/VCR,but Sphase fraction of fas-SGC7901/VCR was about1.9 times that of pBK-SGC7901/VCR,indicatingS phase arrest of fas-SGC7901/VCR.FACS alsosuggested apoptosis of fas-SGC7901/VCR,fas-SGC7901/VCR was more sensitive to apoptosisinducing agent VM-26 than pBK-SGC7901/VCR.MTT assay showed increased sensitization offas-SGC7901/VCR to DDP,MMC and 5-FU,butsame sensitization to VCR according to pBK-SGC7901/VCR.SGC7901,pBK-SGC7901/ VCRand fas-SGC7901/VCR had positively stainedTopo Ⅱ equally.P-gp staining in pBK- SGC7901/VCR was stronger than in SG07901,but there was little staining of P-gp in fas.SGC7901/VCR.CONCLUSION fas gene transduction couldreverse the MDR of human drug-resistant gastriccancer cell SGC7901/VCR to a degree,possiblybecause of higher sensitization to apoptosis anddecreased expression of P-gp.

KAI1 is a potential target for anti-metastasis in pancreatic cancer cells

AIM: To investigate whether KAI1, as a metastasis suppressor gene, is associated with invasive and metastatic ability of pancreatic cancer cells.METHODS: KAI1 gene was transfected into pancreatic cancer cell line MiaPaCa Ⅱ by liposomes selected with G418. Expression of transfected cells was measured by Western blotting, immunofluorescence and immunocytochemistry. Tumor cell invasion and metastatic ability were detected through gelatinase activity and reconstituted basement membrane (Matrigel) assay. pCMV-KAI1 was directly injected into the heterotopic human pancreatic adenocarcinoma successfully established in the groin of BALB/C nude mice, by subcutaneous injection of MiaPaCa Ⅱ pancreatic cancer cells. The statistical analysis between groups was determined by Student's two tailed t test.RESULTS: By Western blotting, MiaPaCa Ⅱ cells transfected by KAI1 gene indicated KAI1 expression at approximately 29.1 kDa. Cytoplasm staining was positive and uniformly spread in transfected cancer cells, using immunohistochemistry and immunofluorescence. The most obvious difference was present after 30 h (MiaPaca Ⅱ 43.6 ± 9.42, pCMV-MiaPaca Ⅱ 44.8 ± 8.56, pCMV-KAI1-MiaPaca Ⅱ 22.0 ± 4.69, P < 0.05). Gelatinolysis revealed a wider and clearer band of gelatinolytic activity in non-transfected than in transfected cells (MiaPaCa Ⅱ cells 30.8 ± 0.57, transfected cells 28.1 ± 0.65, P < 0.05). In vivo tumor growth rates of KAI1 transfectants with KAI1-Lipofectamine 1.22 ± 0.31 in A group were lower than control 4.61 ± 1.98 and pCMV-KAI 11.67 ± 0.81. Analyses of metastases with and without KAI1 transfection in mice were different in liver and lung between controls 1.62 ± 0.39, 0.45 ± 0.09, pCMV-KAI 1.01 ± 0.27, 0.33 ± 0.09 and KAI1-Lipofectamine 0.99 ± 0.21, 0.30 ± 0.09 respectively (P < 0.05).CONCLUSION: High expression of KAI1 gene was found in transfected MiaPaCa Ⅱ human pancreatic cancer cells with lower metastatic ability. KAI1 gene plays an important role in inhibiting metastasis of pancreatic cancer after direct injection into pancreatic adenocarcinoma. These results show that the suppressed invasion and motor function of pancreatic cancer cells may be a key reason why the KAI1 gene controls pancreatic cancer cell metastasis.

In vitro cultivation and differentiation of fetal liver stem cells from mice

During embryonic development, pluripotent endoderm tissue in the developing foregut may adopt pancreatic fate or hepatic fate depending on the activation of key developmental regulators. Transdifferentiation occurs between hepato- cytes and pancreatic cells under specific conditions. Hepatocytes and pancreatic cells have the common endodermal progenitor cells. In this study we isolated hepatic stem/progenitor cells from embryonic day (ED) 12-14 Kun-Ming mice with fluorescence-activated cell sorting (FACS). The cells were cultured under specific conditions. The cultured cells deploy dithizone staining and immunocytochemical staining at the 15th, 30th and 40th day after isolation. The results indicated the presence of insulin-producing cells. When the insulin-producing cells were transplanted into alloxan- induced diabetic mice, the nonfasting blood glucose level was reduced. These results suggested that fetal liver stem/ progenitor cells could be converted into insulin-producing cells under specific culture conditions. Fetal liver stem/ progenitor cells could become the potential source of insulin-producing cells for successful cell transplantation therapy strategies of diabetes.

Localization of AKAP4 and tubulin proteins in sperm with reduced motility

Aim： To perform screening, related to A-kinase anchoring proteins 4 （AKAP4） and tubulin proteins, in spermatozoa with absent or severely reduced motility in order to detect the status of the fibrous sheath and the axonemal structure. Methods： An immunocytochemical study of tubulin, used as a positive control, and AKAP4 was carded out to detect the presence and the distribution of these proteins in different sperm samples. The morphological characteristics of sperm were studied by transmission electron microscope （TEM） and the results were elaborated using a formula reported in previous studies. PCR was carried out on DNA extracted from peripheral blood lymphocytes to analyse partial sequences of the Akap4 and Akap3 genes. Results： Immunolabelling of tubulin and AKAP4 showed different patterns, which led us to divide the patients into groups. In group I, the absence of AKAP4 and tubulin was revealed, although these patients did not show alterations in the Akap4/Akap3 binding site. TEM evaluation highlighted that a high presence of necrosis was associated with total sperm immotility. In group Ⅱ, a regular AKAP4 and tubulin signal was present, although motility was reduced and TEM analysis revealed the presence of immaturity. In group Ⅲ, in which a weak AKAP4 label associated with normal tubulin staining and reduced motility was observed, a severe disorganization of the fibrous sheath was highlighted by TEM. Conclusion： While the role of AKAP4 in sperm motility is unclear, absent or weak AKAP4-1abelling seems to be associated with absent or weak sperm motility.

Aberrant cytological localization of p16 and CDK4 in colorectal epithelia in the normal adenoma carcinoma sequence

AIM： To study the correlation between the patterns of subcellular expression of p16 and CDK4 in colorectal epithelia in the normal-adenoma-carcinoma sequence.METHODS： Paraffin sections of 43 cases of normal colorectal epithelia and corresponding adenomas as well as carcinomas were analysed immunocytochemically for subcellular expression of p16 and CDK4 proteins.RESULTS： Most carcinomas showed more cytoplasmic overexpression for p16 and CDK4 than the adenomas from which they arised or the adjacent normal mucosa. Most normal or non-neoplastic epithelia showed more p16 and CDK4 expression in the nucleus than their adjacent adenomas and carcinomas. There was a significant difference between the subcellular expression pattern of p16 and CDK4 in normal-adenoma-carcinoma sequence epithelia （P 〈 0.001）. Neither p16 nor CDK4 subcellular patterns correlated with histological grade or Dukes＇ stage.CONCLUSION： Interaction of expression of p16 and CDK4 plays an important role in the Rb/p16 pathway.Overexpression of p16 and CDK4 in the cytoplasm, as well as loss expression of p16 in the nucleusmighlc be important in the evolution of colorectal carcinoma from adenoma and, of adenoma from normal epitheiia.

Preparation of monoclonal antibody against human KIAA0100 protein and Northern blot analysis of human KIAA0100 gene

Monoclonal antibodies(MAbs) are important tools for the study of proteins′ function and structure. But there has been no report on the preparation of MAbs against human KIAA0100 protein up to date. Here, first, we generated the mouse MAb against human KIAA0100 protein using purified recombinant 6×Histidinc(6×His)-tagged human KIAA0100 protein segment(1557–2234) as an antigen; then, the m RNA expression of human KIAA0100 gene was detected in U937 cells using Northern blot analysis. The results showed that the mouse MAb against human KIAA0100 protein could sensitively recognize the human KIAA0100 protein using Western blot analysis and immunocytochemistry analysis. Besides, Western blot analysis revealed that human KIAA0100 gene possibly encoded two different protein products(254 k Da and < 250 k Da) in U937 cells. Moreover,Northern blot analysis confirmed that human KIAA0100 gene might produced two different m RNA products(6000–10000 bp and 5000–6000 bp) in U937 cells. The results provide a basis for large-scale production of the MAb against human KIAA0100 protein, which will be useful for the study of human KIAA0100 protein′s function/structure and MAb-targeted drugs in the future.

Intra-operative peritoneal lavage for colorectal cancer

Free cancer cells can be detected in peritoneal fluid at the time of colorectal surgery. Peritoneal lavage in colorectal surgery for cancer is not used in routine, and the prognostic significance of intraperitoneal free cancer cells (IPCC) remains unclear. Data concerning the technique of peritoneal lavage to detect IPCC and its timing regarding colorectal resection are scarce. However, positive IPCC might be the first step of peritoneal spread in colorectal cancers, which could lead to early specific treatments. Because of the important heterogeneity of IPCC determination in reported studies, no treatment have been proposed to patients with positive IPCC. Herein, we provide an overview of IPCC detection and its impact on recurrence and survival, and we suggest further multi-institutional studies to evaluate new treatment strategies.

Occurrence of gonadtropins like substance in the thoracic ganglion mass of the mud crab, Scylla paramamosain (Crustacea: Decapoda: Brachyura)

The identification and localization of vertebrate follicle stimulating hormone （FSH） and luteinizing hormone （LH） in crustaceans may further elucidate the regulation mechanisms in arthropod repro-duction. Using immunocytochemical staining techniques, this study has localized vertebrate FSH-like and LH-like substances in neurons in the subesophageal and thoracic ganglia from the thoracic ganglion mass （TGM） of Scylla paramamosain （Crustacea： Decapoda： Brachyura）. Enzyme-linked immunosorbent assay （ELISA） has shown that the concentrations of both FSH-and LH-like sub-stances increased markedly in the TGM during the vitellogenic stage compared with that in the previtellogenic stage. These results indicate that substances resembling the vertebrate FSH and LH are present in S. paramamosain, and they may be involved in the development of the ovary as well as in ovulation.

Immunocytochemical Identification and Localization of Diffuse Neuroendocrine System (DNES) Cells in Gastrointestinal Tract of Channel Catfish (Ictalurus punctatus)

To detect distribution and relative frequency of diffuse neuroendocrine system （DNES） cells in the gastrointestinal tract of channel catfish （lctalurus punctatus）, the intestinal tract of channel catfish was divided into seven portions from proximal to distal： the enlarged area after oesophagus, cardia, fundus, pylorus, and anterior, middle, and posterior intestine. Immunohistochemical method using the strept avidin-biotin-complex （SABC） was employed. All antisera between seven portions of the channel catfish were compared statistically using statistical package for the social science （SPSS）. Five types of DNES ceils were determined： neuropeptide Y-immunoreactive （NPY-IR） cells were demonstrated in both anterior and middle intestine; serotonin （5-HT） immunoreactive cells were detected throughout the whole gastrointestinal tract; vasoactive intestinal peptide （VIP） positive cells were at the highest frequency in pylorus; glucagon-immunoreactive （GLU-IR） cells were moderate in number in the fundus and anterior, middle intestine, and no immunoreactivity was determined in the other portions; somatostatin （SOM） positive cells were more abundant in the anterior and middle intestine. The regional distribution and relative frequency of immunoreactive cells in the channel catfish, Ictalurus punctatus, are essentially similar to those of other fish. However, some characteristics are observed in this species, which further proved that the diversity of the physiological function of DNES cells was based on their morphology.

Growth associated protein 43 and neurofilament immunolabeling in the transected lumbar spinal cord of lizard indicates limited axonal regeneration

Previous cytological studies on the transected lumbar spinal cord of lizards have shown the presence of differentiating glial cells,few neurons and axons in the bridge region between the proximal and distal stumps of the spinal cord in some cases.A limited number of axons(20-50)can cross the bridge and re-connect the caudal stump of the spinal cord with small neurons located in the rostral stump of the spinal cord.This axonal regeneration appears to be related to the recovery of hind-limb movements after initial paralysis.The present study extends previous studies and shows that after transection of the lumbar spinal cord in lizards,a glial-connective tissue bridge that reconnects the rostral and caudal stumps of the interrupted spinal cord is formed at 11-34 days post-injury.Following an initial paralysis some recovery of hindlimb movements occurs within 1-3 months post-injury.Immunohistochemical and ultrastructural analysis for a growth associated protein 43(GAP-43)of 48-50 k Da shows that sparse GAP-43 positive axons are present in the proximal stump of the spinal cord but their number decreased in the bridge at 11-34 days post-transection.Few immunolabeled axons with a neurofilament protein of 200-220 k Da were seen in the bridge at 11-22 days post-transection but their number increased at 34 days and 3 months post-amputation in lizards that have recovered some hindlimb movements.Numerous neurons in the rostral and caudal stumps of the spinal cord were also labeled for GAP43,a cytoplasmic protein that is trans-located into their axonal growth cones.This indicates that GAP-43 biosynthesis is related to axonal regeneration and sprouting from neurons that were damaged by the transection.Taken together,previous studies that utilized tract-tracing technique to label the present observations confirm that a limited axonal re-connection of the transected spinal cord occurs 1-3 months post-injury in lizards.The few regenerating-sprouting axons within the bridge reconnect the caudal with the rostral stumps of the spinal cord,and likely contribute to activate the neural circuits that sustain the limited but important recovery of hind-limb movements after initial paralysis.The surgical procedures utilized in the study followed the regulations on animal care and experimental procedures under the Italian Guidelines(art.5,DL 116/92).

促生长素抑制素样免疫反应在大白鼠弓状核和正中隆起的超微结构分布

本文用免疫电镜方法证明:促生长素抑制素样免疫反应神经末梢分布于弓状棱并与未标记的树突形成轴树突触。在正中隆起的纤维层和栅状层内均可见上述免疫反应末梢,大多数紧贴门脉毛细血管基底膜周围甚至穿入基底膜内。免疫反应末梢尚可与未标记的末梢形成轴轴突触样结构。

Alteration of chaperonin60 and pancreatic enzyme in pancreatic acinar cell under pathological condition

AIM： To investigate the changes of chaperonin60 （Cpn60） and pancreatic enzymes in pancreatic acinar cells, and to explore their roles in the development of experimental diabetes and acute pancreatitis （AP）. METHODS： Two different pathological models were replicated in Sprague-Dawley rats： streptozotocininduced diabetes and sodium deoxycholate-induced AP. The contents of Cpn60 and pancreatic enzymes in different compartments of the acinar cells were measured by quantitative immunoo/tochemistry. RESULTS： The levels of Cpn60 significanUy increased in diabetes, but decreased in AP, especially in the zymogen granules of the pancreatic acinar cells. The elevation of Cpn60 was accompanied with the increased levels of pancreatic lipase and chymotrypsinogen in diabetes. However, a decreased Cpn60 level was accompanied by high levels of lipase and chymotrypsinogen in AP. The amylase level was markedly reduced in both the pathological conditions. CONCLUSION： The equilibrium between Cpn60 and pancreaUc enzymes in the acinar cells breaks in AP, and Cpn60 content decreases, suggesting an insufficient chaperone capacity. This may promote the aggregation and autoactivation of the premature enzymes in the pancreatic acinar cells and play roles in the development of AR