AIM: To clone and express mouse canstatin (m canstatin)cDNA and provide a basis for the further research on its anti-angiogenic activity and potential application for cancer therapy. METHODS: Total RNA was extracted from mouse liver tissue by Trizol Reagent, and mouse canstatin cDNA was amplified by RT- PCR, then cloned into vector pMD18-T for sequencing. pET30a(+)-m canstatin recombinant plasmid was constructed and expressed in E.coli BL21 with induction of IPTG. RESULTS: Mouse canstatin cDNA is 684 bp coding 227 amino acids. The sequences of both cDNA and amino acid share high homology with human canstatin, with cDNA identity at 89% and amino acids identity at 96% to human canstatin. In the present study, pET30a(+)-m canstatin recombinant plasmid was expressed in E.coli BL21. CONCLUSION: Mouse canstatin cDNA has been cloned for the first time. Constructed pET30a(+)-m canstatin recombinant plasmid is highly expressed in E.coli BL21.

The course of estahlishing coronary collateral circulation has been studied after ligating the anterior descending branch of the left coronary artery (L. A. D.) of the heart in 20 rabbits. Another 5 rabbits were studied as control group. The experimental results were as follows: (1) The rabbit electrocardiograms EGG were made before and after the ligation of the L. A. D. The EGG of control group was made only before killing. In these experiments, the pathological changes of No. 28 were observed immediately after ligating the L. A.D. It was found that the T wave was inverted in standard lead I, and that the volts of the QRS complex in lead Vc fell apparently. This phenomenon may be interpreted as myocardial ischemia in the anterior wall of the heart. After ligating the L. A. D. the S-T segment of No. 27 in AVL lead elevated and exceeded I mm, and the depressive Q wave in Vc lead was greater than the one-fourth of the R wave before killing. All of the phenomenen observed in the rabbit entirely confirmed the myocardial infarction and myocardial ischemia. In No. 25, the T wave was inverted in standard lead I, and the QRS complex in lead Vc was not only shallow in depth but also very little. The conditions accounted probably for myocardial ishcmia. During 4 to 21 days after ligating the L. A. D.,the pathological changes in the EGG occurred frequently.The rabbit ECG gradually became normal from 21 st day on. (2) The coronary arteries of the rabbit hearts were injected with X-ray contrast medium and were examined. The X-ray film thus obtained were compared with the specimen after they were cleared. At the same time, sections of the heart walls (the infarcted area and its surrounding areas) of fifteen rabbits were studied carefully under microscope. Our primary impression was that only a few newly formed vessels in the central part of the infarct till the fourth day after ligating the L. A. D. The vessels surrounding of the infarct show a slight winding and distension. During 10 to 28 days, the number of the newly formed vessels in the infarct increased gradually day by day. The vessels surroundings the infarct were also highly crooked and distended, especially in the anterior wall of the right ventricle. This is possibly the most active period in the establishment of coronary collateral circulation. During 28 to 49 days, the morphology of the vessels in the infarct area became similar to that of the corresponding area of the control group. (3) The significance of the coronary collateral circulation on the heart wall was discussed.

A Study of one hundred hearts (adults 56, children 44) has been made. The S-A mode arteries and A-V node arteries are measured and analysed. The chief results are as fo11ows: (1) 67% of S-A node arteries originated from the right coronary artery; 33% arose from the left coronary artery. A number of the S-A node arteries arising from the right coronary artery were the continuation from the anterior right atrial branch. Occasionally, they may be come from the intermediate right atrial branch or intermediate left atrial branch. (2) The distance between the right S-A node artery origin and the aortic root was about 1.28?0.12 cm in adults, and 0.85?0.09 cm in children, while between the left S-A node artery origin and the bifurcation of the left coronary artery, it was about 0.84?0.19 cm in adults and 0.41?0.03 cm in children. The external diameters of the S-A node arteries were measured. The mean value was as follows: 0.17?0.01 cm (adults); 0.12?0.01 cm (children). (3) The relation between the origins of S-A node artery and A-V node artery may be divided into six types. ①The S-A node artery and A-V node artery which originated from the right coronary artery were about 58.06%. ②The S-A node artery arising from the left coronary artery and the A-V node artery originating from the right coronary artery occurred in 30.11% ③The S-A node artery originated from the right coronary attery, while the A-V node artery arose from the left. This condition was about 6.45%.④The type that the S-A node artery arose from both coronary arteries, and the A-V node artery originated from the right was about 3.22%.⑤Both the S-A node artery and A-V node artery arising from left coronary artery occurred in 1.08%.⑥The A-V node artery originating from the left and right coronary arteries, and the S-A node artery arising from the right coronary artery were about 1.08%.

The coronary arteries of 80 rabbit hearts were studied.Suspension of red lead oxide (Pb_3O_4)in turpentine was injected into the coronary artery.With clearing method and angiographic technique,we studied the branches of the left and right coronary artery, the angle between the anterior descending branch and the left circumflex branch of the left coronary artery,the relationship between the anterior descending artery and the great cardiac vein,the origin of the sinoatrial nodal artery and the atrioventricular nodal artery as well as the type of branch in coronary artery.The results were outlined as follows: 1.The angle between the anterior descending branch and the left circumflex branch is 35～45?in more than one half of the cases(54.5%). 2.The anterior descending branch is often small and short and accompanies the great cardiac vein.It may exist in four forms:1)Both branches run closely to each other. 2)Both run along with each other but leaving a distance in between.3)Both are over- lapping each other.4)Both are crossing each other between the inferior margin of the left auricle and pulmonary artery cone.Ligation of the anterior descending branch in animal experiments should be appropriately made between the right inferior margin of the auricle and the pulmonary artery cone. 3.The left ventricular branch is a direct continuation of the circumflex branch and gives off branches to anterior and posterior wall of the left ventricle. 4.There is an independent ventricular septal artery in the rabbit heart. 5.The sinoatrial nodal artery of the rabbit heart arises frequently from the right coronary artery(86.6%)and the atrioventricular nodal artery from the left coronary artery(72.5%). 6.Commonly the distribution of the coronary blood supply from the left and right coronary arteries to rabbit heart is nearly a balanced circultion(67.5%).Next frequent is the preponderance of the left coronary artery(28.8%),and the preponderance of the right coronary artery occurs only in 3.7%.

Objective To investigate the effects of rapamycin on tumor growth and mTOR/4E-BP1 signaling pathway in xenograft of esophageal squamous cell carcinoma ( ESCC ) EC9706 cells. Methods The expression of factors in mTOR/4E-BP1 signaling pathway of the tumor tissues was analyzed by RT-PCR and Western blot. Results Both rapamycin and cisplatin significantly inhibited the growth of transplantable tumors as compared to control group ( P < 0. 05 ) and their combination had a synergestic effect. The results of RT-PCR and Western blot show that rapamycin significantly decreased the expression of Mtor and p-4E-BP1 but increased the expression of 4E-BP1. Conclusion Rapamycin inhibits the mTOR/4E-BP1 signalling pathway, resulting in the growth inhibition of xenograft of EC9706 cells.

The gross vessels and microvasculature of the rat testis were observed by clear method and corrosion cast method under SEM. The present study noted that there were two kinds of the microvascular arrangement around the circumference of the seminiferous tubule. The first one could be called the intertubular vessels. This vessel was either individual precapillary arteriole or two parallel capillaries connected by a few small branches to form network. The intertubular vessels were located in triangular columns of interstitial tissue between the seminiferous tubules and parallel with the tubules. The second one could be called the peritubular capillaries. These capillaries connected the intertubular vessels in the manner of a rope ladder, and encircled the seminiferous tubule, formed the peritubular capillary network in the subepithelium of the seminiferous tubule. Authors presume that this vascular feature of the architecture surrounding the seminiferous tubules may be related to high blood flow required in the metabolic activity of the tubules and it may contribute to immediate effect of androgen secreting in the interstitial tissue of the testis on the seminiferous tubules. In the article, the gross vascular feature of the arrangement in the rat testis and the microvascular arrangement in the visceral layer of tunica vaginalis of it is described in detail. Their functional significance is also discussed.

Ten fresh eye balls removed from five newborn cadavers were studied.About2-4 hrs after death,cadavers were injected through both common carotid arteries withABS at a constant pressure between 120～150 mmHg.ABS-injected eyes were enucleatedand were cut into halves through equator or meridian.The specimens were digestedin 40% KOH.The corrosion-cast models of blood vessels in uvea were observed underthe SEM.The chief results of observation were as follows:Within the choroid,the blood vessels lie in stratified layers.The large vesselsare located in the outermost layer,next to this layer are smaller vessels.The innermostlayer consists of a continuous single layer of capillaries.The choriocapillaries end around the optic disc.This finding indicates that thechoriocapillaries are not directly involved in the disc circulation.Each short posterior ciliary artery supplied a segment of the choriocapillaries,with some capillary anastomosis between segments,so that the choriocapillaries werearranged in one plane as a single continuous layer of capillaries forming a network.The network of the choriocapillaries consisted of lobules of choriocapillaries.The precapillary arteriole lie in its center.In each lobule meshed capillaries appearedto radiate from a central focus.The postcapillary venule lay on the periphery of thelobule.The lobules varied in their geometric configuration,forming irregularlytriangular to hexagonal shapes.The average size of a lobules of newborn's chorioca-pillaries was 230/420? to 460/520?.The diameter of the choriocapillary varies between8? and 20?.The blood vessels of the ciliary body and iris divide from large circular arteryof iris and long posterior ciliary arteries.Each ciliary process contained a small arterywhich randomly branched to form a capillary network.It is drained by venule.

The architecture of the alveolus and its capillaries of human lung injected withsolution of ABS in methyl ethyl ketone was studied under the SEM.The results wereoutlined as follows:1.The alveolar casts were observed from the subpleural and intralobular septu-lar surfaces.The human alveolus is irregular polygon in appearance.The size ofthe alveolus is variable,its surface is smooth,there are many depressions of the topof type Ⅱ cells.The bridge-like structure between two adjacent alveoli are thecasts of the alveolar pores.They are variable both in size and number,and ofround or oval shapes with smooth surface.2.Capillaries of the subpleural space and interlobular septulum are transitional,and identical in appearance.Meshes of the capillary network are larger than thoseof other parts,but more closer in density compared with those in dog.Each capil-lary is branched from the metaarteriole.3.Capillaries in the alveolar septum is a single layer of dense network,theirdiameter are larger than those of the mesh hole.They originate mainly from thecapillaries of subpleural space and interlobular septulum.

OBJECTIVETo explore the differences in sensitivity to rapamycin of five esophageal squamous cell carcinoma cell lines with different differentiation and the changes of sensitivity of the cells after siRNA-interfered expression of p70S6K.

METHODSEffects of rapamycin on proliferation of ESCC cell lines with different differentiation, EC9706, TE-1, Eca109, KYSE790 and KYSE450 cells, were investigated using cell counting kit 8 (CCK-8) assay, and according to the above results, the EC9706 cells non-sensitive to rapamycin were chosen to be transfected with p70S6K-siRNA. The changes in sensitivity of cells to rapamycin were measured in vitro and in vivo using CCK-8 kit, flow cytometry and tumor formation in nude mice.

RESULTSCCK-8 results showed that all the five cell line cells were sensitive to low concentration of rapamycin (<100 nmol/L), but TE-1 and EC9706 cells, which were with poor differentiation, showed resistance to high concentration of rapamycin. After EC9706 cells were treated with 50, 100, 200, 500 and 1 000 nmol/L rapamycin and p70S6K-siRNA, the proliferation rates of EC9706 cells were (48.67 ± 1.68)%, (15.45 ± 1.54)%, (14.00 ± 0.91)%, (10.97 ± 0.72)% and (2.70 ± 0.32)%, respectively, and were significantly lower than that of cells treated with 50, 100, 200, 500 and 1 000 nmol/L rapamycin and control siRNA [(74.53 ± 1.71)%, (68.27 ± 1.35)%, (71.74 ± 2.44)%, (76.23 ± 1.02)% and (80.21 ± 2.77)%] (P<0.05 for all). The results of flow cytometry showed that the ratios of cells in G1 phase of the p70S6K-siRNA, rapamycin and p70S6K-siRNA+ rapamycin groups were (53.82 ± 1.78)%, (57.87 ± 4.01)% and (73.73 ± 3.68)%, respectively, significantly higher than that in the control group (46.09 ± 2.31)% (P<0.05 for all). The results of tumor formation test in vivo showed that the inhibitory effect of rapamycin on tumor growth was stronger after the cells were transfected with p70S6K-siRNA, and the inhibition rate was 96.5%.

CONCLUSIONESCC cells with different differentiation have different sensitivity to rapamycin, and p70S6K-siRNA can improve the sensitivity of cells to rapamycin in vitro and in vivo.

Animals ; Antibiotics, Antineoplastic ; pharmacology ; Carcinoma, Squamous Cell ; drug therapy ; metabolism ; pathology ; Cell Differentiation ; Cell Line, Tumor ; Cell Proliferation ; Esophageal Neoplasms ; drug therapy ; metabolism ; pathology ; Humans ; Mice ; Mice, Nude ; RNA, Small Interfering ; Ribosomal Protein S6 Kinases, 70-kDa ; genetics ; metabolism ; Signal Transduction ; Sirolimus ; pharmacology ; Transfection