Experimental study on diabetic cardiomyopathy rats treated with NMFGF1 loaded PEG-modified nano-liposomes combined with ultrasound-targeted microbubble destruction technique / 中华内分泌代谢杂志

Objective To investigate the therapeutic effect and mechanism of non-mitogenic acid fibroblast growth factor 1( NMFGF1) on diabetic cardiomyopathy ( DCM) by using PEG-modified nano-liposomes combined with ultrasound-targeted microbubble destruction technique ( UTMD ) . Methods The NMFGF1 loaded PEG-modified nano-liposomes were prepared by a water-in-water emulsion method and their quality inspections were also investigated. Type 1 diabetes animal model was induced by intraperitoneal injection of streptozotocin ( 70 mg/kg) in male SD rats. The diabetic rats were raised twelve weeks after the diabetes model was established and DCM rats were selected by ultrasonic heart function examination. After two weeks of intervention, all rats were kept for another two weeks and then underwent transthoracic echocardiography examination. The rats were sacrificed and myocardial tissue was obtained to quantify myocardial collagen fraction ( CVF ) and cardiac myocyte apoptotic index by Sirius red staining and TUNEL staining. Results NMFGF1-loaded PEG-nano-liposomes showed a good morphology and 90.3％± 1.4％ NMFGF1 encapsulation efficiency. Compared with DCM group, NMFGF1group, and NMFGF1-PEG-nano-liposomes group, NMFGF1-loaded PEG-nano-liposome plus UTMD group showed increased left ventricular end diastolic diameter (LVIDd) [(7.36±0.42) vs (5.75±0.24), (6.64±0.27), (6.72±0.24)mm, all P<0.05]and leftventricularfractionshortening(LVFS) [(50±3) vs (33±2), (44±5), (43±3)mm, all P<0.05], and decreased left ventricular posterior wall thickness (LVPW) [(1.65±0.07) vs (1.89±0.08), (1.73±0.11), (1.73 ±0.07) mm, all P<0.05], with decreased CVF and apoptotic index(all P<0.05). Conclusion PEG-nano-liposomes combining with UTMD technique has a greater translational potential in the delivery of NMFGF1 for the treatment of DCM by attenuating oxidative stress-induced injury and may provide a promising strategy for treating diabetes cardiomyopathy.

Snake venom galactoside-binding lectins: a structural and functional overview

Snake venom galactoside-binding lectins (SVgalLs) comprise a class of toxins capable of recognizing and interacting with terminal galactoside residues of glycans. In the past 35 years, since the first report on the purification of thrombolectin from Bothrops atrox snake venom, several SVgalLs from Viperidae and Elapidae snake families have been described, as has progressive improvement in the investigation of structural/functional aspects of these lectins. Moreover, the advances of techniques applied in protein-carbohydrate recognition have provided important approaches in order to screen for possible biological targets. The present review describes the efforts over the past 35 years to elucidate SVgalLs, highlighting their structure and carbohydrate recognition function involved in envenomation pathophysiology and potential biomedical applications.

Snake venom galactoside-binding lectins: a structural and functional overview

Snake venom galactoside-binding lectins (SVgalLs) comprise a class of toxins capable of recognizing and interacting with terminal galactoside residues of glycans. In the past 35 years, since the first report on the purification of thrombolectin from Bothrops atrox snake venom, several SVgalLs from Viperidae and Elapidae snake families have been described, as has progressive improvement in the investigation of structural/functional aspects of these lectins. Moreover, the advances of techniques applied in protein-carbohydrate recognition have provided important approaches in order to screen for possible biological targets. The present review describes the efforts over the past 35 years to elucidate SVgalLs, highlighting their structure and carbohydrate recognition function involved in envenomation pathophysiology and potential biomedical applications.(AU)

Molecular Characterization and Mitogenic Activity of a Lectin from Purse Crab Philyra Pisum

A lectin from the hemolymph of purse crab, Philyra pisum, was found to have anti-proliferative activity on human lung cancer cells by our laboratory. In this study, P. pisum lectin (PPL) was molecularly characterized including molecular mass, amino acid sequences, amino acid composition, and the effects of metal ions, temperature, and pH on the activity. We found that PPL showed mitogenic activity on human lymphocytes and BALB/c mouse splenocytes. The mitogenic activity (maximum stimulation index, SI=9.57+/-0.59) of PPL on human lymphocytes was higher than that of a standard well-known plant mitogen, concanavalin A (maximum SI=8.80+/-0.59). The mitogenic activity mediated by PPL is required for optimum dosing, and higher or lower concentrations caused decreases in mitogenic response. PPL also induced mitogenic activity on mouse splenocytes, however, the maximum SI (1.77+/-0.09) on mouse splenocytes of PPL was lower than that (2.14+/-0.15) of concanavalin A. In conclusion, PPL is a metal ion-dependent monomer lectin with mitogenic activity, and could be used as a lymphocyte or splenocyte stimulator.

Papel dos fatores de crescimento na regeneração periodontal: estado atual da arte / The role of growth factors in periodontal regeneration: current state-of-the-art

Fatores de crescimento polipeptídicos são proteínas de baixo peso molecular sintetizadas por várias células e tecidos que regulam os eventos básicos que levam à regeneração periodontal: proliferação, quimiotaxia, diferenciação e síntese de matriz protéica. Embora diversos métodos terapêuticos tenham sido propostos para uso clínico visando a regeneração dos tecidos periodontais, a previsibilidade de sucesso ainda é limitada, com casos bem-sucedidos e outros mostrando resultados aquém do esperado. Recentemente, após a identificação da existência de células tronco no ligamento periodontal e a compreensão que a diferenciação destas células em outras mais diferenciadas (fibroblastos, osteoblastos e cementoblastos) é mediada por moléculas de sinalização (fatores de crescimento), alguns estudos foram publicados propondo a utilização clínica dos fatores de crescimento, isolados ou combinados, em diferentes tipos de veículos com o objetivo de favorecer a regeneração periodontal. Neste artigo, as características gerais e os estudos in vitro e in vivo evidenciando o papel dos principais fatores de crescimento utilizados na regeneração periodontal serão revistos com o objetivo de avaliar o papel dos fatores de crescimento na regeneração periodontal.

Polipeptidic growth factors are low density proteins secreted by different cells and tissues that regulate the basic events leading to periodontal regeneration: proliferation, chemotaxis, differentiation and protein synthesis. Although many methods for achieving regeneration of lost periodontal tissues have been suggested, there is poor predictability, with some well-succeeded cases and others showing unexpected results. Recently, after the identification of mesenchymal stem cells in the periodontal ligament and the understanding that the differentiation of these cells onto more differentiated ones (osteoblasts, cementoblasts, fibroblasts) is mediated by signaling molecules (growth factors), some studies have proposed the clinical use of growth factors isolated or in combination to enhance periodontal regeneration outcomes. In this paper, the characteristics and in vitro and the in vivo studies evaluating the role of growth factors in periodontal regeneration will be reviewed to evaluate the role of growth factors in periodontal regeneration.

Expression and Purification of Basic Fibroblast Growth Factor Mutant with Reduced Mitogenic Activity / 中国生物工程杂志

In order to decrease the potential side-effects of human basic fibroblast growth factor (hbFGF) caused by its broadspectrum mitogenic activity, a single residue of hbFGF, the residue serine 108, was replaced with neutral alanine residue to construct a mutant of hbFGF (mhbFGF) with reduced mitogenic activity. The mutant was overexpressed in Escherichia coli BL21(DE3) by IPTG induction. The expression level of mhbFGF was about 30% of the total cellular protein. The expressed mhbFGF was purified by ionic exchange and heparin affinity chromatography from the supernatant of bacteria lysate. Measured by MTT method, the effect of mhbFGF on Balb/c 3T3 cell proliferation was much lower than that of wild-type hbFGF. The purified recombinant mhbFGF was prepared and sufficient for the following pharmacological and safety studies.

Expression and Purification of Basic Fibroblast Growth Factor Mutant with Reduced Mitogenic Activity / 中国生物工程杂志

In order to decrease the potential side-effects of human basic fibroblast growth factor (hbFGF) caused by its broadspectrum mitogenic activity, a single residue of hbFGF, the residue serine 108, was replaced with neutral alanine residue to construct a mutant of hbFGF (mhbFGF) with reduced mitogenic activity. The mutant was overexpressed in Escherichia coli BL21(DE3) by IPTG induction. The expression level of mhbFGF was about 30% of the total cellular protein. The expressed mhbFGF was purified by ionic exchange and heparin affinity chromatography from the supernatant of bacteria lysate. Measured by MTT method, the effect of mhbFGF on Balb/c 3T3 cell proliferation was much lower than that of wild-type hbFGF. The purified recombinant mhbFGF was prepared and sufficient for the following pharmacological and safety studies.

Expression and Purification of Basic Fibroblast Growth Factor Mutant with Reduced Mitogenic Activity / 中国生物工程杂志

In order to decrease the potential side-effects of human basic fibroblast growth factor (hbFGF) caused by its broadspectrum mitogenic activity, a single residue of hbFGF, the residue serine 108, was replaced with neutral alanine residue to construct a mutant of hbFGF (mhbFGF) with reduced mitogenic activity. The mutant was overexpressed in Escherichia coli BL21(DE3) by IPTG induction. The expression level of mhbFGF was about 30% of the total cellular protein. The expressed mhbFGF was purified by ionic exchange and heparin affinity chromatography from the supernatant of bacteria lysate. Measured by MTT method, the effect of mhbFGF on Balb/c 3T3 cell proliferation was much lower than that of wild-type hbFGF. The purified recombinant mhbFGF was prepared and sufficient for the following pharmacological and safety studies.

Comparison between Immunostimulatory Activity and Molecular Structure of Different Polysaccharides

BACKGROUND: We previously reported that ginsan, a polysaccharide extracted from Panax ginseng had an immunostimulatory activity such as mitogenic activity, activation of macrophages and killer cells, and production of a variety of cytokines which resulted in antitumor and antiseptic effects. We further purified alpha-(1-->6)-glucan and beta-(2-->6)-fructan from the ginsan with size exclusion and ion-exchange column chromatography successively. In this study, we performed the structure-based activity of ginsan by comparison with known polysacchrides such as beta-glucan, curdlan, laminarin, levan, dextran, lentinan and OK-432. METHODS: To investigate the immunostimulatory activity of several polysaccharide compounds, we investigated the stimulation of lymphocytes proliferation, the generation of activated killer cells and the secretion of nitrites from activated macrophages. RESULTS: Of polysaccharides tested, curdlan and ginsan stimulated lymphocyte proliferation, suggesting that the molecular weight and composition of polysaccharide are dependent on the mitogenic activity. The production of nitric oxide was significantly increased in curdlan, levan, ginsan and its fraction, indicating that fructan has also capacity to activate macrophages and may devote to kill pathogens. In addition, the activation of macrophages was seemed to be independent of molecular weight of polysaccharide. The generation of AK cells was exhibited in order of curdlan, OK-432> F1, ginsan, F3>levan>etc. The AK activity may be dependent on molecular weight and composition of polysaccharides. CONCLUSION: Unfortunately, purified polysaccharide from ginsan were less active on immunostimulatory activity than mixed compounds of polysaccharides. From the viewpoint of structure and activity relationships, we found several characteristic features.

B radykinin Receptor-Meadiated Intracellular Calcium Mobilization and ATP Changes in Cultured Bovine Corneal Endothelial Cell

To clarify the effect of bradykinin(Bk)on cultured bovine corneal endothelial cells(BCEC), cytosolic free calcium([Ca2+])mobilization and cell proliferation were investigated. The [Ca2+] was determined using a Ca2+ sensitive indicator, Fura-2/AM, and cell proliferation was evaluated by counting the cell number. Bk induced the transient increase of [Ca2+] in a concentration-dependent manner(10(-11)M~10(-7)M)and its 50% effective concentration was about 5x10(-11)M. The basal [Ca2+] with 1mM CaCl2 in the bathing solution was 87+/-9nM. Transient Bk(10(-8)M)-induced [Ca2+] increase was inhibited slightly but significantly by the pretreatment with EGTA. The pretreatment with U-73122(5x10(-6)M), an inhibitor of phospholipase C, also attenuated Bkinduced [Ca2+] mobilization. To identify and characterize the Bk receptor subtype in BCEC, Bk1 and Bk2 antagonists were investigated. Transient Bk(10(-8)M)-induced [Ca2+] increase was almost absolutely attenuated by the pretreatment with Bk2 antagonist for 10 minutes. To investigate the physiological effect of Bk, Bk-induced mitogenic effect was studied. 10(-8)M of Bk produced significant increase of intracellular ATP levels from the day 2 to the day six of culture period. This Bk-induced mitogenic effect was inhibited by the treatment with Bk2 antagonist. Bk-induced ion transport was determined by measuring intracellular ATP contents. Intracellular ATP content([ATP]i)was decreased by the treatment with 10(-8)M Bk for 10 minutes. Bk-induced [ATP]i decrement was significantly restored by the pretreatment with ouabain for 30 minutes. In summary, stimulation of intracellular signal transduction by Bk in BCEC is coupled with Bk2 type receptor. And also, Bk produces mitogenic effect and enhancesion and fluid transport in BCEC.

Mitogenic effects of nicotine to human periodontal ligament(PDL) cells in vitro / 대한치과교정학회지

Nicotine is one of the major components of cigafette smoking which causes various systemic and local diseases to human body. Mitrogenic effects of nicotine to systemic disease are interesting factors in the results of cellular proliferation especially to vascular and pulminary tissus or cells. The study of local effects concerns with destruction of tissue and delayed healing rate after various surgicla treatment. Platelet-Derived Growth factor(PDGF) and Insulin-like growth factor(IGF) are known as major mitogens to human PDL cells. The purpose of this studywas to investgate the mitogenic effects of nicotine to human PDL cells. We studied the expression of PDGF-alpha receptor, PDGF-betareceptor, and IGF-1 receptor mRNA form the nicotine treated human PDL cells by northern analysis. The experimental groups were divided into different serum(1%, 10%) and nicotine(100ng/ml, 1000ng/ml) condentrations and each group was studied by time course. The results of this study showed upregulation of PDGF-alpha, beta receptor and IGF-1 receptor mRNA at 100ng/ml nicotine concentration and 10% serum group to the time course. These results suggest that physiologically attainable nicotine concentrations may stimulate mitogenic gene synthesis to human PDL cells in vitro.

Corneal neovascularization induced by controlled-release of interleukin-2 in rabbits / 眼科

Corneal neovascularization(CNV)is one of the main causes of corneal allograft rejection.We have developed a CNV model in rabbits by controlled- release of IL-2.Its characterization and developmental processes were observed and its histological mechanism was explored.CNV was revealed through staining of erythrocytes within capillaries with 3,3′-diaminoben- zidine(DAB).Ethylene vinyl acetate copolymer (EVA-40)pellets incorporated with IL-2 100u,200u, 500u and control blank pellets were implanted in corneal mid-stromal pocket 2mm from the upper limbus.Biomicroscopic evaluation revealed that IL-2 induced CNV in a dose-dependent manner.IL-2 500u induced CNV in all the experimental eyes with good consistency and reproducibility.Histopathologi- cally,the IL-2 induced CNV was closely associated with mononuclear cell infiltration.This model fairly closely mimicked the immunologically mediated CNV. The architecture and perfusion of CNV was clearly revealed by the red blood cells stained with DAB.

Studies on Lymphocyte Subpopulations and Cell-mediated lmmunity in Patients with Chronic Renal Failure

Several parameters of cell-mediated immunity thirty-eight patients with end stage chronic renal failure were measured including total lymphyocytes, B-and T-lymphocytes, T-cell subsets and the mitogenic reponse to PHA and Con A at three different times; before dialysis, 3 months and 12 months after dialysis treatment. There were no significant differences in the absolute numbers of peripheral leukocytes between each patient and the control group. But the absolute numbers of lymphocytes of each patient group were significantly reduced compared to the control group (p< 0.01). The proportion of peripheral blood active T cells and helper T cells was significantly reduced both in the predialysis uremic and dialysis populations compared to the control group, although the helper/suppressor(OKT4/OKT8) ratio was not different between each patient and the control group except for a lower ratio in the hemodialysis 12 month follow-up group (HD 12M). With respect m the PHA and Con A stimulation tests, the stimulation indices of the predialysis and hemodialysis groups were significantly lower than those of the control group. However, patients on continuous ambulatory peritoneal dialysis (CAPD) exhibited a normal mitogenic response and a lower suppressor cell removal index compared to the patients on hemodialysis, suggesting an improved cell-mediated immunity in the patients undergoing CAPD.

Protective effect of non-mitogenic haFGF on cerebral ischemia-reperfusion injury in mice / 中国药理学通报

Aim To investigate the protective effect of non-mitogenic human acidic fibroblast growth factor (nm-haFGF) on cerebral ischemia-reperfusion injury in mice. Methods Cerebral ischemia-reperfusion model was made by ligating bilateral carotid for 20 minutes in mice. These mice were randomly divided into model group( iv NS), two doses of nm-haFGF (iv 25、50 ?g?kg-1) groups, rhaFGF group(iv 50 ?g?kg-1) and sham- operated group. Step down test and Y-type electric maze were used to examine the effect of nm-haFGF on learning and memory of mice, then Even′s Blue(EB) level and NO level in brain of these mice were measured. Results The nm-haFGF significantly decreased numbers of errors of mice in 5 min in step down test and in Y-type electric maze test; EB and NO levels in brain of these mice were lower than those of model group respectively. Conclusion The nm-haFGF can protect cerebral ischemia-reperfusion injury in mice.

In vitro study of the effect of selenium on normal human T-lymphocyte functions / 中国病理生理杂志

The effect of selenium on T cell PHA response, on the IL2 productionand on the proliferative response of lymphocytes to IL-2 was studied on in vitro normalblood. Lymphocytes of normal peripheral blood cultured in the presence of sodium sele-nite showed increased proliferation in response to PHA and produced more IL-2 thanthose cultured with PHA alone. But the proliferative response of lymphoblasts to IL-2was not effected by sodium selenite.

A study on status of the immunity in alloxan diabetic mice and the immunoregulation of insulin / 中国病理生理杂志

This paper report the effects of hyperglucosemia and hypoinsulinemia on Immune system of alloxan diabetic mice. The results showed that the weight of thymus and spleen as well as the YAC-1 cell cleaning rate in the organs (lung, liver, blood) of the diabetic mice were significantly lower than that of the control. The proliferation response to Con A(2.5-20?g/ml) and the production of IL-2 of the lymphocytes were inhibi-ted markedly. When the lymphocytes were suspended in the culture medium containiny insulin, the proliferation response and the production of IL-2 increased markedly. These results suggested that the functions of lymphocytes and their natural killer activity were impaired in diabetic mice. Insulin is one of the important immunoregulation hormones and plays an important role in the regulation of immune system.

Influence of head x-irradiation on reactivity of lymphocytes to Con A and IL-2 production in rat spleen / 中国病理生理杂志

The proliferative response of splenic lymphocytes to Con A in Wistar rat was significantly enhenced after x-irradiation of the head with 10 Gy. At a cell concentration of 5?10~6 per ml, the ratio of 3~H-TdR incorporation between the irradiated head group and whole-body shielded group was 3.20(P

Hydrocortisone inhibited ConA-activated thymus cell IL-2 restoration test / 中国病理生理杂志

IL-2 plays an important role in immune response regulation. Detection and quantitation of IL-2 is based on the use of an IL-2 dependent T cell (CTLL) or the IL-2 dependent proliferation of PHA-activated blasts. Both such clones are difficult to obtain and maintain. IL-2 assay on thymocyte activity was also used but it lacks of specificity because it also responses to IL-1.Christine (1984) developed a simple method for detection of IL-2. This is adopted in this work with some modification, and the result is compared with that obtained from CTLL assay.In brief, the ConA-activated(4ug/ml)proliferation of C57BL/6 mouse thymus cell suspension is completely blocked by hydrocortisone (HC 0.4ug/ml) The inhibition was restored by IL-2 but not by IL-1. the highest dilution of the IL-2, which restored the activity of the ConA-activated HC blocked thymus cells, is used as the titer of IL-2 quantitation for a given specimen. This was compared with that of IL-2 which can activate the CTLL. A close correlation between the titers obtained by these two assays. (r=0.94) However, the sensitivity of CTLL assay is slightly higher than that with HCIT(0.4IL-2 u and1.3 IL-2 u respectively).Hence, this technique for IL-2 detection is specific as to IL-1 is concerned, mocre feasible than the current CTLL assay, and relatively inexpensive.

Postburn Kinetic Changes of Stress Hormones and Splenocyte Immune Functions in Rats / 第三军医大学学报

After the major burn a multitude of immunologic alterations develop. Although the roles of suppressor cells and serum inhibitory factors in the mechanism of postburn immunosuppression are well known, the role of the neuroendo-crine response is still not understood. In this paper determining stress hormones and splenocyte immune functions are simultaneously focused on in deeply burned rats with 25% BSA. The changes of plasma corticosterone (B) and catecholamine are dominant within 24 h postburn(PB), and soon return to normal. The endogenous activity of splenocytes (SBT) decreases at 12 h PB. while the ability of spleno-cytes responsing to ConA in vitro (MSBT) is almost unchanged. The pathological lesions of spleen may be affected by increasing levels of plasma B. When most of the stress hormones return to normal at 72 h PB the SBT and MSBT become different. This may be influenced by many other factors rather than the neuroendocrine system. The differences between SBT and MSBT suggest thai SBT may reflect the more real condition of lymphocytes in vivo than MSBT in vitro. Following burns the roles of streess hormones actually participate in the immune regulation such as plasma B and catecholamine act in the early postburn stage.