Purification and chemical modification of porcine bone morphogenetic protein

Implantation of porcine bone morphogenetic protein (pBMP) in the muscle induces differentiation of mesenchymal-type cells and results in endochondral bone formation. pBMP was isolated from porcine demineralized bone matrix and purified by hydroxyapatite chromatography, Sephadex G75 gel filtration, preparative sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), preparative isoelectric focusing (IEF), and chromatofocusing fast protein liquid chromatography (FPLC). Porcine BMP has an MW of 26 K and a range of pI from 4.65 to 4.73 determined by SDS-PAGE and IEF, respectively. Reconstitution with the citrate buffer supernatant fraction enables as little as 50 micrograms of the soluble pBMP fractions to induce osteogenesis in an in vivo assay. Chemical modification studies indicate that the osteoinductive potential of the pBMP molecule depends on tyrosine, carboxyl groups, and disulfide bonds and can be increased by modification of sulfhydryl groups. Modification of arginine and tryptophan has no effect on bioactivity. By pepsin-limited proteolysis, fragments of pBMP with an MW of 6-14 K show definite, although reduced, BMP activity.     

烧伤大鼠血清对血管内皮细胞通透性的影响

加深入阐明严重烧伤早期全身血管通透性增加继而引直民早期损害的发生机制，本实验应用培养于微孔滤膜上的小牛瞌 及皮细胞单层所构成的选择性通透屏障模型，观察了３０％Ⅲ度烧伤大鼠血清对其影响，结果发现，烧伤早期烧伤血清刺激后ＥＣ单层对Ｈａｎｋｓ液或白蛋白液滤过流量、滤过系数均明显增加。     

大剂量卡铂化疗的药物毒性观察

6例肺癌患者接受了大剂量卡铂并自体骨髓移植治疗 ,卡铂剂量从 5 60～ 1375 mg/ m2 ,5例加用 VP～ 1630 0 mg,1例并用 MMC6mg和 VCR2 mg;主要毒性反应为白细胞、中性粒细胞、血小板减少和脱发 ( WHO毒性反应 ～ 级 )其次是贫血、皮肤粘膜出血、呕吐和可逆性肝功能损害 ,口腔粘膜溃疡及轻度发烧各 2例 ,一过性蛋白尿、室上性心动过速和便秘各 1例 ,尿素氮和肌酐未见升高 ;全部患者骨髓均获解救 ,从自体骨髓回输到骨髓造血功能重建成功平均 2 6.67( 17～ 32 )天     

Three years' experience with levonorgestrel-releasing intrauterine device and Norplant-2 implants: a randomized comparative study

A randomized comparative study of levonorgestrel-releasing intrauterine device (LNG-IUD) and Norplant^R-2 implants was carried out in 200 women for 36 months. Three thousand one hundred woman-months of use were observed with each device. Only one pregnancy occurred in users of LNG-IUD at the 12th month of use. The change in bleeding pattern was the most frequent reason for discontinuation. The discontinuation rate for irregular bleeding with Norplant-2 was 17.3 and 26.8 at 24 and 36 months, respectively, as compared to 3.3 with LNG-IUD at both 24 and 36 months. The differences were statistically significant. Removal for amenorrhea and pain only occurred in acceptors of LNG-IUD. About 20–40% of women using Norplant-2 had prolonged bleeding through 36 months. The percentage of amenorrhea in LNG-IUD was the highest (29.3%) at the end of two years of use. More than 97% of subjects reported satisfaction with the methods used by themselves.

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Resumen Un estudio aleatorizado en el que se comparan los dispositivos intrauterinos que descargan levonorgestrel (LNG-IUD) con los implantes Norplant-2 se realizó en 200 mujeres durante 36 meses. Se observó un período de utilización correspondiente a tres mil cien meses-mujer con cada uno de los dispositivos. Hubo un solo embarazo, en el decimosegundo mes, entre las usuarias de LNG-IUD. La alteración del flujo menstrual fue la razón invocada con mayor frecuencia para el abandono del método. Los porcentajes de abandono por la irregularidad de la menstruación en el caso de los Norplant-2 fue de 17,3 y 26,8 a los 24 y 36 meses, respectivamente, en comparación con 3,3 con los LNG-IUD a los 24 y 36 meses. Estas diferencias son estadísticamente significativas. El retiro por amenorrea y dolores sólo ocurrió entre las mujeres que tenían los LNG-IUD. Los Norplant-2 ocasionaron sangrado prolongado durante los 36 meses estudiados en alrededor del 20 al 40% de las mujeres. El porcentaje de amenorrea con los LNG-IUD llegó al nivel máximo (29,3%) al cabo de dos años de uso. Más del 97% de las mujeres se mostraron satisfechas con los métodos que utilizaban.

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Resumé Une étude randomisée comparant les dispositifs intra-utûrins libérant du lévonorgestrel (LNG-IUD) aux implants Norplant-2 a été effectuée sur 200 femmes pendant 36 mois. On a ainsi observé une période d'utilisation correspondant à trois mille cent mois/femme avec chacun des dispositifs. Une seule grossesse s'est produite, au douzième mois, parmi les utilisatrices de LNG-IUD. L'altération du flux menstruel a été la raison le plus souvent invoquée pour l'abandon de la méthode. Les pourcentages d'abandon du fait de l'irrégularité de la menstruation dans le cas des implants se sont élevés à 17,3 et 26,8 après respectivement 24 et 36 mois, alors qu'ils n'atteignaient que 3,3 pour les LNG-IUD, tant à 24 qu'à 36 mois. Ces différences sont statistiquement significatives. Le retrait à la suite d'aménorrhée et de douleurs n'a été constaté que chez des femmes portant les LNG-IUD. Le Norplant-2 a occasionné des saignements prolongés pendant les 36 mois étudiés chez environ 20 à 40% des femmes. Dans le cas des LNG-IUD, le taux d'aménorrhée était le plus élevé (29,3%) au terme de deux années d'utilisation. Plus de 97% des patientes se sont dites satisfaites de la méthode qu'elles appliquaient.

     

肢体不同延长速度对局部血流量的影响

本文报告20只兔胫骨以不同速度延长时用末梢微循环测定系统动态连续监测局部血流量变化的结果,发现一次延长>1 mm时,局部肌肉血流量开始急骤下降,恢复延长前血流量所需的时间>8 h。作者认为肢体的血管、肌肉等组织对1 mm/次/d延长速度有相当大的生物适应性。延长速度>1 mm/次/d易造成局部血流量大幅度减少。延长肢肌肉萎缩、肌力减弱可能与血流量减少有关。     

Changes in hippocampal neurons and memory function during the developmental stage of newborn rats with hypoxic-ischemic encephalopathy

BACKGROUND: Under the normal circumstance, there exist some synapses with inactive functions in central nervous system (CNS), but these functions are activated following nerve injury. At the early stage of brain injury, the abnormal functions of brain are varied, and they have very strong plasticity and are corrected easily. OBJECTIVE: To observe the changes of neuronal morphology in hippocampal CA1 region and memory function in newborn rats with hypoxic-ischemic encephalopathy(HIE) from ischemia 6 hours to adult. DESIGN: Completely randomized grouping, controlled experiment. SETTING: Taian Health Center for Women and Children; Taishan Medical College. MATERIALS: Altogether 120 seven-day-old Wistar rats, of clean grade, were provided by the Experimental Animal Center, Shandong University of Traditional Chinese Medicine. Synaptophysin (SYN) polyclonal antibody was provided by Maixin Biological Company, Fuzhou. METHODS: This experiment was carried out in the Laboratory of Morphology, Taishan Medical College between October 2000 and December 2003. ① The newborn rats were randomly divided into 2 groups: model group and control group, 60 rats in each group. Five rats were chosen from each group at postoperative 6 hours, 24 hours, 72 hours, 7 days, 2 weeks and 3 weeks separately for immunohistochemical staining. Fifteen newborn rats were chosen from each group at postoperative 4 weeks and 2 months separately for testing memory ability (After test, 5 rats from each group were sacrificed and used for immunohistochemical staining)② The right common carotid artery of newborn rats of model group was ligated under the anesthetized status. After two hours of incubation, the rats were placed for 2 hours in a container filled with nitrogen oxygen atmosphere containing 0.08 volume fraction of oxygen, thus, HIE models were created; As for the newborn rats in the control group, only blood vessels were isolated, and they were not ligated and hypoxia-treated. ③ Thalamencephal tissue sections of newborn rats of two groups were performed DAB developing and haematoxylin slight staining. Cells with normal nucleous in 250 μm-long granular layer which started from hippocampal CA1 region were counted with image analysis system under high-fold optical microscope (×600), and the thickness of granular layer was measured. The absorbance (A) of positive reactant of SYN in immunohistochemically-stained CA1 region was measured. Learning and memory ability were measured with step through test 3 times successively. ④ t test and paired t test were used for comparing intergroup and intragroup difference of measurement data respectively, and Chi-square for comparing the difference of enumeration data. MAIN OUTCOME MEASURES: Comparison of cytological changes in hippocampal CA1 region and memory ability at different postoperative time points between two groups. RESULTS: Totally 120 newborn rats were involved in the result analysis. ① Cell morphological changes in hippocampal CA1 region: In the control group, with aging, perikaryon, nucleus and nucleolus in cortex of parietal lobe were significantly increased, Nissl body was compacted, the amount of neurons was declined, but the A of SYN positive reactant was relatively increased. In the model group, at postoperative each time point, neurons were seriously shrunk and dark-stained, nucleus was contracted, chromatin was condensed, nucleolus was unclear, even cells disappeared, especially the cells in 6 hours and 24 hours groups. The amount of neurons with normal morphology in hippocampal CA1 region and granular layer thickness in the model group at postoperative each time point were significantly less or smaller than those in the control group at postoperative 6 hours respectively (t =3.002-1.254, P < 0.01). The A value of SYN positive reactant at postoperative 2, 3 and 4 weeks was significantly higher than that at previous time point (t =2.011－2.716,P < 0.05－0.01). ② Test results of learning and memory ability: In the first test, there was no significant difference in the ratio of rats which kept memory ability between two groups (P > 0.05); In the third test, the ratio of rats which kept memory ability in the model group was significantly lower than that in the control group at postoperative 4 weeks and 2 months[53%(8/15)，100%(15/15)；60%(9/15)，93%(14/15)，χ 2=2.863，2.901，P < 0.01]. CONCLUSION: The destroyed hippocampal structure induces the decrease of learning and memory ability of developmental rats. Early interference can increase the quality of neurons and also promote functional development of the nervous system.     

Experimental scoliosis in the rat. II. Biomechanical analysis of the forces during Harrington distraction

The purpose of this study was to characterize the forces resulting from Harrington distraction of the spine in an experimental model of scoliosis in the rat, in order to establish both the similarity of this model to human scoliosis and identify potential force parameters that may be useful for clinical decision-making. Harrington distraction was performed in 36 rats that had been made scoliotic 9-12 weeks earlier by the method described in the previous paper. Distractions were carried out in discrete and timed steps until separation of the vertebral laminae (mechanical failure) occurred at the upper hook site. Distractive forces were monitored continuously by a strain gauge mounted on the tension side of the upper arm of the outrigger. The resulting data were compared among the various curvature groups. The relationship between the length of distraction and the maximum force produced was similar for all animals regardless of curvature. This relationship was quadratic and was characterized by an inflection point where forces increased rapidly with each distraction. The amount of distraction necessary to reach both the inflection and failure points differed only for curves above 100 degrees. The amount of force required to reach failure was lower for curves above 75 degrees. Curves above 50 degrees had a lower percent correction at the inflection point. Bending and tensile forces were calculated by vector analysis. Axial load efficiencies were greater for curves above 50 degrees, as evidenced by increased bending forces in these animals. The viscoelasticity of the spine decreased after inflection in all animals.(ABSTRACT TRUNCATED AT 250 WORDS)     

A cocaine analog and a GBR analog label the same protein in rat striatal membranes.

Because some evidence suggests that cocaine and GBR12935 bind to different sites, we utilized photoaffinity probes from both classes of compounds to see if they label the same protein. [125I]RTI-82 a cocaine analog, and [125I]DEEP, a GBR analog, labeled protein(s) showing the same molecular weight, a similar pharmacological profile and a similar sensitivity to neuraminidase.     

Repression of the embryonic myosin heavy chain phenotype in regenerating chicken slow muscle is dependent on innervation.

Ventricular-like and fast myosin heavy chains (VL-MHC and FMHC) are transiently expressed during slow skeletal muscle development. The influence of innervation on repression of these MHC isoforms is investigated over an 84-day time course in: (1) normal anterior latissimus dorsi (N-ALD) muscles, (2) regenerating ALD (R-ALD) muscles, (3) denervated ALD (D-ALD) muscles, and (4) regenerating and denervated ALD (RD-ALD) muscles. Western blotting demonstrates that the VL-MHC is expressed in R-, D-, and RD-ALD muscles, but not in N-ALD muscles. Expression of the VL-MHC is transient in R-ALD muscles. In contrast, VL-MHC expression persists in RD-ALD muscles, and appears with time in D-ALD muscles. FMHC was not detected in N-ALD muscles by Western blotting. Two FMHCs are seen in R-ALD and RD-ALD muscles, and in 13-day embryonic ALD muscles. The slower migrating FMHC (FMHCA) comigrates with developmentally regulated FMHCs in fast pectoralis muscle, while the faster migrating FMHC (FMHCB) comigrates with the faster migrating FMHC in embryonic ALD muscle (13 days in ovo). FMHCB decreases in amount over the time course in R-ALD muscles, while FMHCA persists. In contrast, substantial levels of both FMHCs persist in RD-ALD muscles, and appear with time in D-ALD muscles. The cellular distribution of MHCs is followed by immunocytochemistry. Regenerating cells expressing VL-MHC and FMHC are replaced by a mature population in R-ALD muscles. Some of the mature myofibers in R-ALD muscles express FMHC, but not VL-MHC. In RD-ALD and D-ALD muscles, both regenerating and mature muscle cells are seen which express VL-MHC and FMHC. Our results indicate that innervation is required for the repression of VL-MHC and FMHCB during regeneration of slow muscle.     

Transitional mucosa at anastomosis. A cause of local tumor recurrence in patients with rectal cancer after anterior resection.

Transitional mucosa adjacent to colorectal cancers is essentially characterized by an excess of sialomucins at the expense of the normally predominant sulphomucins in epithelial cells lining the intestinal crypts which presents the early stage of oncogenic transformation of colorectal epithelium. The presence or absence of sialomucins at the resection margins was studied histochemically using the high iron diamine-alcian blue(HID-AB) stain in 64 rectal cancer patients in Dukes' B stage who underwent curative anterior resection. The correlation was revealed between the presence of sialomucins at the resection margins and subsequent development of local tumour recurrence. Fourteen of 27 patients (51.9%) with sialomucins predominant pattern at either resection margin developed local recurrence compared with 4 of 37 patients (10.8%) with sulphomucins predominant pattern (P less than 0.001). It is suggested that determination of the transitional mucosa around anastomosis in patients treated for the rectal carcinoma by anterior resection appears to identify those with a higher risk of local recurrence.