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Objective To investigate the effect of Lumbricus protein on the phenotypic transformation of corporal cavernosum smooth muscle cells(CCSMC)and erectile function in diabetic erectile dysfunction(DMED)rats.Methods Sixty male SD rats with normal erectile function were randomly divided into a blank group,a model group,a Sildelafil group(5 mg·kg-1),and a Lumbricus protein low-,medium-,and high-dose group(45,90,and 180 mg·kg-1),with 10 rats in each group.The diabetic rat model was established by intraperitoneal injection of Streptozotocin(STZ,50 mg·kg-1)combined with high-fat feed feeding;after 8 weeks,the DMED rat model was prepared by neck injection of Apomorphine(APO,100 μg·kg-1).After successful modeling,the rats were administered with a dose of Apomorphine by gavage once a day for 4 weeks.The blood glucose levels and body mass of rats in each group were measured before modeling,on the third day of modeling,and after 4 weeks of drug administration.The intracavernous pressure(ICP)and carotid artery pressure(MAP)were measured by multi-channel physiological recorder,and the ICP/MAP ratio was calculated.The expressions of contractile markers α-smooth muscle actin(α-SMA),smooth muscle myosin heavy chain(SMMHC)and synthetic markers Collagen I and osteopontin(OPN)in corpus cavernosum were detected by immunohistochemistry.The mRNA expression levels of α-SMA,SMMHC and Collagen I in corpus cavernosum were detected by RT-PCR.The protein expression levels of α-SMA,Desmin,Collagen I and OPN in corpus cavernosum were detected by Western Blot.Results Compared with the blank group,the blood glucose levels of the rats in the model group were significantly increased on the third day of modeling and after 4 weeks of administration(P＜0.01),and the body mass was significantly decreased after 4 weeks of administration(P＜0.01).ICP and ICP/MAP ratio were significantly decreased(P＜0.01).The protein expression levels of α-SMA,SMMHC and Desmin in penile corpus cavernosum were significantly decreased(P＜0.01),and the protein expression levels of Collagen I and OPN were significantly increased(P＜0.01).The mRNA expression levels of α-SMA and SMMHC in corpus cavernosum were significantly decreased(P＜0.01),and the mRNA expression level of Collagen I was significantly increased(P＜0.01).Compared with the model group,there was no significant change in blood glucose and body mass of rats in the administration group(P＞0.05).ICP and ICP/MAP ratio were significantly increased(P＜0.01).The expression levels of α-SMA,SMMHC and Desmin in corpus cavernosum were significantly increased(P＜0.01),while the expression levels of Collagen I and OPN were significantly decreased(P＜0.01).The mRNA expression levels of α-SMA and SMMHC in corpus cavernosum were significantly increased(P＜0.01),and the mRNA expression level of Collagen I was significantly decreased(P＜0.01).Conclusion Lumbricus protein can improve the erectile function of DMED rats,and its mechanism may be related to the inhibition of CCSMC from'contractile'to'synthetic(proliferative)'transformation.
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@# <b>Objective:</b>to observe the effect and mechanism of lumbricus peptides on early renal injury in spontaneous hypertensive rats (SHR) based on Toll-like receptors 4 (TLR4)/nuclear factor-<i>κ</i>B(NF-<i>κ</i>B) signaling pathway. <b>Method:</b>The 40 SHRs were randomly divided into model group (equal volume of distilled water by intragastric administration), lumbricus peptides low, middle and high dose groups (126, 252, 504 mg·kg<sup>-1</sup>), and Benazepril group (0.9 mg·kg<sup>-1</sup>), <i>n</i>=8 in each group. 8 male rats with normal blood pressure at the same age were set as the normal control group,with equal volume of distilled water. After treatment for 60 consecutive days, enzyme-linked immunosorbent assay (ELISA) was used to determine microalbumin (mAlb)and <i>N</i>-acetyl-<i>β</i>-<i>D</i>-glucosaminidase (NAG) content in 24 h urine as well as the level of serum angiotensinⅡ (AngⅡ). Ultrastructural changes of rat kidneys were observed by transmission electron microscope. Western blot was used to detect renal TLR4, NF-<i>κ</i>B p65 protein levels. Immunohistochemical method was used to detect renal tumor necrosis factor-<i>α</i> (TNF-<i>α</i>) and interleukin 10 (IL-10) expression. <b>Result:</b>As compared with the normal control group, the levels of urine mAlb, NAG and serum Ang Ⅱ were increased in the model group (<i>P</i><0.05); the expression of TLR4 and NF-<i>κ</i>B p65 protein was increased (<i>P</i><0.05); expression of TNF-<i>α</i> was increased (<i>P</i><0.05), while IL-10 expression was decreased (<i>P</i><0.05). Transmission electron microscopy of kidney tissues showed that most of the glomeruli of the model group had podocyte foot process fusion, mesangial cells and mesangial matrix hyperplasia. As compared with the model group, the levels of urine mAlb, NAG, and serum Ang Ⅱ were decreased in the rats in lumbricus peptides groups and benazepril group (<i>P</i><0.05); the expression of TLR4 and NF-<i>κ</i>B p65 protein was decreased (<i>P</i><0.05); the positive expression of TNF-<i>α</i> in kidney was decreased to different extent (<i>P</i><0.05), but the expression of IL-10 was increased (<i>P</i><0.05). Transmission electron microscopy of kidney tissues showed that the damage of kidneys in rats after administration of high-dose lumbricus peptides and benazepril was improved in varying degrees. <b>Conclusion:</b>lumbricus peptides can reduce early renal damage in SHRs, and its mechanism may be achieved by regulating the AngⅡ-TLR4/NF-<i>κ</i>B pathway.
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Objective@#To observe the expression of alpha smooth muscle actin (α-SMA) and high mo-bility group protein B1 (HMGB1) in silicosis model rats interfered by lumbricus.@*Methods@#45 rats were ran-domly divided into the control group, model group and group interfered by lumbricus. The silicosis model rats were established. The group interfered by lumbricus were intragastric administered with lumbricus decoction by the 4 ml/kg dose. The control group and model group were ig administered with the equal amount of normal saline. Each group were killed 5 rats on the 7th, 14th and 28th day. The lung tissues were stained with HE and Sirius red methods. The mRNA expressions of α-SMA and HMGB1 were determined with RT-PCR; The pro-tein levels of α-SMA and HMGB1 were determined with Western blotting.@*Results@#Compared with the control group, the expression levels of α-SMA and HMGB1mRNA and protein in lung tissue of model group were grad-ually increased in the 7th, 14th and 28th days, the difference was statistically significant (P< 0.01) . Compared with model group, the levels of α-SMA and HMGB1mRNA and protein in lung tissue of group interfered by lumbricus were gradually lowered in the 7th, 14th and 28th days, the difference was statistically significant (P<0.05, P<0.01) .@*Conclusion@#Lumbricus inhibits the collagen deposition and the formation of silicosis pulmo-nary fibrosis, which may be related to the inhibition of HMGB1 expression and activation of α-SMA in lung tis-sue.
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On the basis of pre-experiment research and the hypothesis of“amputated lumbricus”, this research was aimed to explore mechanism of active components of the amputated lumbricus to promote wound healing. Skin excision was used to establish the mice model. The amputated lumbricus extract was prepared. HE staining and immunohistochemistry techniques were used in the determination of the wound healing rate and changes of VEGF, bFGF, TGF-β1 expression during wound healing period. The results showed that compared with the blank control group, the healing rate of the amputated lumbricus extract group was better. And the HE staining showed better improvement of traumatic tissues. There was no statistic differences on the expression of VEGF and TGF-β1 between the amputated lumbricus extract group and the normal saline group (P> 0.05). The expression of bFGF in amputated lumbricus extract group reached peak earlier than the control group and also lasted a longer time. The amputated lumbricus extract group reached peak on the first day, which had a significant difference (P < 0.05) compared with the control group at the same timepoint. It was concluded that the external application of amputated lumbricus extract had wound healing effect on traumatic skin of mice. Its mechanism may be irrelevant to the expression of VEGF and TGF-β1. However, it may be related to the increasing of bFGF expression in the injured regions during the inflammation stage and proliferation stage.
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Objective: To optimize the technology of freeze-dried powder of various fractions from lumbricus (Pheretima) by Box-Behnken design and response surface method. Methods: Using single factor experiment with Box-Behnken response surface method, freeze-dried rate as the index, the effects of sublimation temperature, sublimation time, analytical drying temperature, and drying time on the freeze-drying process were observed; The freeze-drying process of the active components in lumbricus was optimized; Using HPLC method, the fingerprint of the sample by the optimized technology was investigated, and the similarity of fingerprint was compared with the earthworm medicine, and the optimum freeze-drying process was obtained. Results: The best freeze-drying process conditions of active components in lumbricus were as follows: frozen temperature of -26.5 ℃, precool for 4 h, sublimation drying temperature of -20 ℃, sublimation time of 7 h, analytical drying temperature of 30 ℃, analytical drying time of 3.5 h, freeze-dried rate of 96.55%, and compared with medicinal materials of amino acid, the similarities of fingerprint were both greater than 0.9. Conclusion: Box-Behnken response surface method is used for the freeze-drying process conditions of active components in the lumbricus, the optimization is feasible, the effect of model is good, and this optimization process has the feasibility.
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Objective To investigate the mechanisms of Chinese traditional medicine mixture protection of vascular endothelial cell from apoptosis in cerebral ischemia-reperfusion injury. Methods Healthy, clean SD rats were randomly divided into 4 groups: Sham opera-tion group (SOG), cerebral ischemia reperfusion injury group (IRG), cerebral ischemia preconditioning group (IPG) and Chinese tradi-tional medicine mixture preconditioning group (CPG). Furthermore, IRG, IPG and NPC were divided into 4 sub-groups: 1 d, 7d, 14d, 21d subgroup, according to the different time point since ischemia-reperfusion took place. And in CPG, Naotai formula extract was used. Cere-bral vascular endothelial cells of rats were removed and Hoechst 33258 staining and the DNA gradient bands were used to detect the apoptosis of these cells. Then the influence of Naotai formula extract on caspase-3, 8 and 9 activation, and Bid lysis was examined by Western-blot, and the mechanisms of Chinese traditional medicine mixture protection of vascular endothelial cell were investigated from apoptosis signal pathway. Result Naotai formula extract can inhibit the apoptosis of endothelial cells in ischemia-reperfusion injury, and it also can inhibit the activation of caspase-3, 8 and 9, thus inhibit the lysis of Bid into tBid. Conclusion Naomi formula extract inhibit the apoptosis of endo-thelial ceils in ischemia-reperfusion injury via apoptosis signal pathway.
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Objective To study partial biochemical characterization and effect of lumbricus dornases (LDs). Methods Sephacryl-200 column chromatography, CIEF and agarose electrophoresis were used to determine LDs. Results The molecular weights of LD1 and LD2 were 126 kDa and 62 kDa respectively. The pI of LD1 and LD2 were 6.12 and 6.05. LDs degraded bacterial plasmid and ?DNA. Conclusion LDs is a new kind of dornases. The determination of molecular weights and pI of LDs will help their further study. The results of plasmid and ?DNA degradation imply that LDs can oppose bacteria drug resistance.
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Objective To study preliminar il y the mutual antigens between Lumbricus terrestris[WT5”BZ ] and Schistosoma japonicum. Methods The antigen s of Lumbricus terrestris (L.terrestris) and [ WT5”BX]Schistosoma japonicu m (S.japonicum) were analysed by SDS-PAGE technique, and reacte d with the sera of patients with schistosomiasis and the sera of rabbits immunized with LtAg by Dot-ELISA and Western-blot. Results [WT5” B Z]There were many similar molecular weight antigens between LtAg and SjAg. Spe cially some antigens between 30-90 kDa form LtAg had intensive cross reactio n with the sera of patients with schistosomiasis. SjAg was also reacted with the sera of r abbits immunized with LtAg. Conclusion There are at least three immunocompetent antigens with similar molecular weight between L.terrestris and S.japonicum. [