Noninvasive Electrical Stimulation Improves Photoreceptor Survival and Retinal Function in Mice with Inherited Photoreceptor Degeneration.

Purpose: Neurons carry electrical signals and communicate via electrical activities. The therapeutic potential of electrical stimulation (ES) for the nervous system, including the retina, through improvement of cell survival and function has been noted. Here we investigated the neuroprotective and regenerative potential of ES in a mouse model of inherited retinal degeneration. Methods: Rhodopsin-deficient (Rho-/-) mice received one or two sessions of transpalpebral ES or sham treatments for 7 consecutive days. Intraperitoneal injection of 5-ethynyl-2'-deoxyuridine was used to label proliferating cells. Weekly electroretinograms were performed to monitor retinal function. Retinal morphology, photoreceptor survival, and regeneration were evaluated in vivo using immunohistochemistry and genetic fate-mapping techniques. Müller cell (MC) cultures were employed to further define the optimal conditions of ES application. Results: Noninvasive transpalpebral ES in Rho-/- mice improved photoreceptor survival and electroretinography function in vivo. ES also triggered residential retinal progenitor-like cells such as MCs to reenter the cell cycle, possibly producing new photoreceptors, as shown by immunohistochemistry and genetic fate-mapping techniques. ES directly stimulated cell proliferation and the expression of progenitor cell markers in MC cultures, at least partially through bFGF signaling. Conclusions: Our study showed that transpalpebral ES improved photoreceptor survival and retinal function and induced the proliferation, probably photoreceptor regeneration, of MCs; this occurs via stimulation of the bFGF pathways. These results suggest the exciting possibility of applying noninvasive ES as a versatile tool for preventing photoreceptor loss and mobilizing endogenous progenitors for reversing vision loss in patients with photoreceptor degeneration.

Catharmus tinctorius volatile oil promote the migration of mesenchymal stem cells via ROCK2/Myosin light chain signaling.

MSC transplantation has been explored as a new clinical approach to stem cell-based therapies for bone diseases in regenerative medicine due to their osteogenic capability. However, only a small population of implanted MSC could successfully reach the injured areas. Therefore, enhancing MSC migration could be a beneficial strategy to improve the therapeutic potential of cell transplantation. Catharmus tinctorius volatile oil (CTVO) was found to facilitate MSC migration. Further exploration of the underlying molecular mechanism participating in the pro-migratory ability may provide a novel strategy to improve MSC transplantation efficacy. This study indicated that CTVO promotes MSC migration through enhancing ROCK2 mRNA and protein expressions. MSC migration induced by CTVO was blunted by ROCK2 inhibitor, which also decreased myosin light chain (MLC) phosphorylation. Meanwhile, the siRNA for ROCK2 inhibited the effect of CTVO on MSC migration ability and attenuated MLC phosphorylation, suggesting that CTVO may promote BMSC migration via the ROCK2/MLC signaling. Taken together, this study indicates that C. tinctorius volatile oil could enhance MSC migration via ROCK2/MLC signaling in vitro. C. tinctorius volatile oil-targeted therapy could be a beneficial strategy to improve the therapeutic potential of cell transplantation for bone diseases in regenerative medicine.

Eradication Efficacy of Modified Dual Therapy Compared with Bismuth-Containing Quadruple Therapy as a First-Line Treatment of Helicobacter pylori.

OBJECTIVES: This study assessed the effectiveness, adverse events, patient adherence, and costs of modified dual therapy compared with bismuth-containing quadruple therapy for treating Helicobacter pylori infection in Chinese patients. We also sought to determine whether modified dual therapy could be used as an alternative first-line treatment for H. pylori infection. METHODS: A total of 232 H. pylori-infected, treatment-naive patients were enrolled in this open-label, randomized controlled clinical trial. Patients were randomly allocated into 2 groups: the 14-day modified dual therapy group and the bismuth-containing quadruple therapy group. Eradication rates, drug-related adverse events, patient compliance, and drug costs were compared between the 2 groups. RESULTS: The modified dual therapy group achieved eradication rates of 87.9%, 91.1%, and 91.1% as determined by the intention-to-treat, per-protocol, and modified intention-to-treat analyses, respectively. The eradication rates were similar compared with the bismuth-containing quadruple therapy group: 89.7%, 91.2%, and 90.4%. In addition, modified dual therapy ameliorated variations in the CYP2C19, IL-1B-511, and H. pylori VacA genotypes. There were no significant differences in the compliance rates between the 2 groups. The modified dual therapy group exhibited significantly less overall side effects compared with the bismuth-containing quadruple therapy group (P < 0.001). Furthermore, the cost of medications in the modified dual therapy was lower compared with that in the bismuth-containing quadruple therapy. CONCLUSIONS: Modified dual therapy at high dose and administration frequency is equally effective and safer and less costly compared with bismuth-containing quadruple therapy.

(+)-Cholesten-3-one induces osteogenic differentiation of bone marrow mesenchymal stem cells by activating vitamin D receptor.

In our previous reports, it was revealed that steroids in traditional Chinese medicine (TCM) have the therapeutic potential to treat bone disease. In the present study, an in vitro model of a vitamin D receptor response element (VDRE) reporter gene assay in mesenchymal stem cells (MSCs) was used to identify steroids that enhanced osteogenic differentiation of MSCs. (+)-cholesten-3-one (CN), which possesses a ketone group that is modified in cholesterol and cholesterol myristate, effectively promoted the activity of the VDRE promoter. Phenotypic cellular analysis indicated that CN induced differentiation of MSCs into osteogenic cells and increased expression of specific osteogenesis markers, including alkaline phosphatase, collagen II and Runt-related transcription factor 2. Furthermore, CN significantly increased the expression of osteopontin, the target of the vitamin D receptor (VDR), which indicated that CN may activate vitamin D receptor signaling. Over-expression of VDR or knockdown studies with VDR-small interfering RNA revealed that the pro-differentiation effects induced by CN required VDR. Furthermore, the present study determined that the C-terminal region of the VDR is responsible for the action of CN. Taken together, the present findings demonstrated that CN induced osteogenic differentiation of MSCs by activating VDR. The present study explored the regulation of stem cells by using a series of similar steroids and provided evidence to support a potential strategy for the screening of novel drugs to treat bone disease in the future.

Intravenous Esomeprazole for Prevention of Peptic Ulcer Rebleeding: A Randomized Trial in Chinese Patients.

INTRODUCTION: High-dose intravenous esomeprazole is the only approved pharmacological treatment for the prevention of peptic ulcer rebleeding (currently approved in over 100 countries worldwide), but has not yet been approved in China. This study aimed to evaluate a high-dose esomeprazole intravenous regimen vs. an active control (cimetidine) for the prevention of rebleeding in Chinese patients with a high risk of peptic ulcer rebleeding who had undergone primary endoscopic hemostatic treatment. METHODS: This was a parallel-group study conducted at 20 centers in China. The study comprised a randomized, double-blind, intravenous treatment phase of 72 h in which 215 patients received either high-dose esomeprazole (80 mg + 8 mg/h) or cimetidine (200 mg + 60 mg/h), followed by an open-label oral treatment phase in which all patients received esomeprazole 40 mg tablets once daily for 27 days. The primary outcome was the rate of clinically significant rebleeding within the first 72 h after initial endoscopic hemostatic therapy. Secondary outcomes included the rates of clinically significant rebleeding within 7 and 30 days; proportions of patients who had endoscopic retreatment and other surgery due to rebleeding; and number of blood units transfused. RESULTS: The rate of clinically significant rebleeding within 72 h was low overall (3.3%) and numerically lower in patients treated with esomeprazole compared with cimetidine (0.9% vs. 5.6%). Overall, the results of the secondary outcomes also showed a numerical trend towards superiority of esomeprazole over cimetidine. All treatments were well tolerated. CONCLUSION: In this phase 3, multicenter, randomized trial conducted in China, esomeprazole showed a numerical trend towards superior clinical benefit over cimetidine in the prevention of rebleeding in patients who had successfully undergone initial hemostatic therapy of a bleeding peptic ulcer, with a similar safety and tolerability profile. These findings suggest that esomeprazole may be an alternative treatment option to cimetidine for this indication in China. FUNDING: AstraZeneca. TRIAL REGISTRATION: ClinicalTrials.gov identifier, NCT01757275.

[Effect of retinociacdi combined extracts from testudinis carapacis et plastri on proliferating in MSCs and its mechanism].

OBJECTIVE: To explore the effect of retinociacdi (RA) combined extracts from Testudinis Carapacis et Plastri(PTE) on proliferating in MSCs and its mechanism. METHODS: Transfected PGL3-ID1 using the calcium phosphate co-precipitation method in rat MSCs. PTE combined with RA and retinociacdi receptor inhibitor(Ro41) acted on transfected MSCs with respective concentrations of 10(-6), 10(-7) and 10(-8) mol/L. Luciferase activity measurement was used to detect the activity of RAR and IDI 36 h later. PTE acted on MSCs 36 h,3 d and 7 d for respective concentrations of 1, 3, 30 and 100 microg/mL,then collected cells to detect RAR with RT-PCR. PTE combined with RA for 10(-7) mol/L and Ro41 for 10(-6) mol/L respectively on MSCs for 36 h,and then collected cells to detect RAR and ID1 with RT-PCR. RESULTS: PTE promoted expression of ID1 on MSCs. When combined with RA, the promotion effect became greater and it promoted expression of RAR at the same time; When inhibited RA using Ro41, the promotion of IDI was weaken by PTE. CONCLUSION: RA promotes expression of IDI on MSCs, PTE regulates proliferation and differentiation of MSCs by expression of nuclear receptor RAR.

Polyethylene Glycol Electrolyte Lavage Solution versus Colonic Hydrotherapy for Bowel Preparation before Colonoscopy: A Single Center, Randomized, and Controlled Study.

This single center, randomized, and controlled study aimed to compare the effectiveness and safety of polyethylene glycol electrolyte lavage (PEG-EL) solution and colonic hydrotherapy (CHT) for bowel preparation before colonoscopy. A total of 196 eligible outpatients scheduled for diagnostic colonoscopy were randomly assigned to the PEG-EL (n = 102) or CHT (n = 94) groups. Primary outcome measures included colonic cleanliness and adverse effects. Secondary outcome measures were patient satisfaction and preference, colonoscopic findings, ileocecal arrival rate, examiner satisfaction, and cecal intubation time. The results show that PEG-EL group was associated with significantly better colonic cleanliness than CHT group, fewer adverse effects, and increased examiner satisfaction. However, the CHT group had higher patient satisfaction and higher diverticulosis detection rates. Moreover, the results showed the same ileocecal arrival rate and patient preference between the two groups (P > 0.05). These findings indicate that PEG-EL is the preferred option in patients who followed the preparation instructions completely.

ß-Asarone prevents autophagy and synaptic loss by reducing ROCK expression in asenescence-accelerated prone 8 mice.

ß-Asarone is an active component of the Acori graminei rhizome that is a traditional Chinese medicine clinically used in treating dementia in China. However, the cognitive effect of ß-asarone and its mechanism has remained elusive. Here, we used asenescence-accelerated prone 8 (SAMP8) mice, which mimic many of the salient features of Alzheimer׳s disease (AD), to further investigate whether modulation of the ROCK signaling pathway and/or autophagy, synaptic loss is involved in the effects of ß-asarone on learning and memory. SAMP8 mice at the age of 6 months were intragastrically administered by ß-asarone or a vehicle daily for 2 months. Senescence-accelerated-resistant (SAMR1) mice were used as the control. Our results demonstrate that autophagy and ROCK expression were increased significantly in 8 months SAMP8 mice, which were concomitant with that SAMP8 mice at the same age displayed a significant synaptic loss and cognitive deficits. The up-regulation of ROCK expression and autophage in the hippocampus of SAMP8 were significantly reduced by ß-asarone, and prevents synaptic loss and improved cognitive function of the SAMP8 mice. ß-asarone decreased neuronophagia and lipofuscin in the hippocampus of SAMP8 mice, but did not reduce Aß42 levels and malondialdehyde levels and superoxide dismutase activities. Moreover, suppression of ROCK2 by siRNA significantly reduced the effects of ß-asarone on the autophage and synaptic proteins expression in PC12 cells damage induced by Aß1-40. Taken together, ß-asarone prevents autophagy and synaptic loss by reducing ROCK expression in SAMP8 mice.

Muscone exerts neuroprotection in an experimental model of stroke via inhibition of the fas pathway.

Identifying small molecules that are neuroprotective against stroke injury will be highly beneficial for treatment therapies. A cell viability assay and gas chromatography-mass spectrometry were used to identify active small molecules in XingNaoJing, which is a well known Chinese medicine prescribed for the effective treatment of stroke. Studies have found that muscone is the active compound that prevents PC12 cell and cortical neuron damage following various injuries. Analysis of apoptosis indicated that muscone inhibited glutamate-induced apoptotic cell death of PC12 cells and cortical neurons. Fas and caspase-8 expression were upregulated following glutamate treatment in cortical neurons, and was markedly attenuated in the presence of muscone. Furthermore, muscone significantly reduced cerebral infarct volume, neurological dysfunction and inhibited cortical neuron apoptosis in middle cerebral artery occluded (MCAO) rats in a dose-dependent manner. Moreover, a significant decrease in Fas and caspase-8 expression in the rat cortex was observed in MCAO rats treated with muscone. Our results demonstrate that muscone may be a small active molecule with neuroprotective properties, and that inhibition of apoptosis and Fas is an important mechanism of neuroprotection by muscone. These findings suggest a potential therapeutic role for muscone in the treatment of stroke.

Targeting of the Sonic Hedgehog pathway by atractylenolides promotes chondrogenic differentiation of mesenchymal stem cells.

Molecules that enhance chondrogenic differentiation in mesenchymal stem cells (MSCs) were identified and isolated using an in vitro Gli reporter gene assay in MSCs incorporating a Sonic Hedgehog (Shh) target. Atractylenolide III, which promoted Gli1-mediated transcriptional activity, was isolated from an ethyl acetate extract of the Rhizoma, Atractylodis macrocephalae. After dehydration, atractylenolide III was transformed to atractylenolide I. Both atractylenolides were confirmed by MS, UV, IR, 1H- and 13C-NMR spectra. Atractylenolide III (which contains -OH at the 8-position) and atractylenolide I (which lacks -OH at the 8-position) were found to effectively promote the activity of the Gli promoter. While the hydroxyl group of atractylenolide III was not essential for the effect of atractylenolide, its effect was dependent on Shh signaling. Phenotypic cellular analysis indicated that atractylenolides induced MSCs to differentiate into chondrocytes, as shown by increased expression of specific chondrogenic markers including collagen II, aggrecan and the cartilage related transcription factor, Sox9. Atractylenolides significantly increased the expression of Shh and its target gene Gli-1, indicating that Shh signaling was activated by atractylenolides. Moreover, inhibition of Shh signaling reduced the effect of atractylenolides on the chondrogenic phenotype. The discovery that atractylenolides induce chondrocytes from MSCs is promising for bony disease therapy.

Microbead-induced ocular hypertensive mouse model for screening and testing of aqueous production suppressants for glaucoma.

PURPOSE: To characterize the microbead-induced ocular hypertension (OHT) mouse model and investigate its potential use for preclinical screening and evaluation of ocular hypotensive agents, we tested the model's responses to major antiglaucoma drugs. METHODS: Adult C57BL/6J mice were induced to develop OHT unilaterally by intracameral injection of microbeads. The effects of the most commonly used ocular hypotensive drugs, including timolol, brimonidine, brinzolamide, pilocarpine, and latanoprost, on IOP and glaucomatous neural damage were evaluated. Degeneration of retinal ganglion cells (RGCs) and optic nerve axons were quantitatively assessed using immunofluorescence labeling and histochemistry. Thickness of the ganglion cell complex (GCC) was also assessed with spectral-domain optical coherence tomography (SD-OCT). RESULTS: A microbead-induced OHT model promptly responded to drugs, such as timolol, brimonidine, and brinzolamide, that lower IOP through suppressing aqueous humor production and showed improved RGC and axon survival as compared to vehicle controls. Accordingly, SD-OCT detected significantly less reduction of GCC thickness in mice treated with all three aqueous production suppressants as compared to the vehicle contol-treated group. In contrast, drugs that increase aqueous outflow, such as pilocarpine and latanoprost, failed to decrease IOP in the microbead-induced OHT mice. CONCLUSIONS: Microbead-induced OHT mice carry dysfunctional aqueous outflow facility and therefore offer a unique model that allows selective screening of aqueous production suppressant antiglaucoma drugs or for studying the mechanisms regulating aqueous humor production. Our data set the stage for using GCC thickness assessed by SD-OCT as an imaging biomarker for noninvasive tracking of neuronal benefits of glaucoma therapy in this model.

Chemical components in extracts from Plastrum testudinis with proliferation-promoting effects on rat mesenchymal stem cells.

Plastrum Testudinis (PT) is often used as an important traditional Chinese medicine to treat bone diseases in China for centuries. To identify active components of PT involved in promoting proliferation of MSCs, PT was extracted with ethyl acetate and separated by silica gel column chromatography with gradient elution. Sixteen (Ts-1 ≈ 16) components were obtained, which were biologically evaluated by MTT assay and flow cytometry on the proliferation of rat marrow-derived mesenchymal stem cells (rMSCs). Results indicated that Ts-12 could induce the proliferation compared with control group (p < 0.05), while Ts-4 exhibited inhibitive effect. The chemical components of PT which regulated the proliferation of rMSCs were investigated by Gas Chromatography-Mass Spectrometry (GC-MS), HPLC, and nine standard compounds. The experimental results suggested that palmitic acid methyl ester and cholesterol myristate, which identified from Ts-12, possessed proliferative activity while stearic acid found in Ts-4 showed inhibition.

[Active ingredients of Plastrum Testudinis inhibit epidermal stem cell apoptosis in serum-deprived culture].

OBJECTIVE: To investigate the effects of active ingredients of Plastrum Testudinis (PT) on serum deprivation-induced apoptosis of epidermal stem cells (ESCs). METHODS: ESCs were isolated from the back skin of fetal Sprague-Dawley rats with 2 weeks of gestational age and were divided into normal group (10% fetal bovine serum), control group (serum-deprived culture) and groups treated with serum deprivation plus active ingredients of PT, including ethyl acetate extract (2B), stearic acid ethyl ester (S6), tetradecanoic acid sterol ester (S8) and (+)-4-cholesten-3-one (S9). The vitality of ESCs after 24, 48 and 72 h of culture was measured with MTT method; apoptotic ESCs double-stained with Annexin V-FITC and propidium iodine were detected by flow cytometry (FCM); Bcl-2 and caspase-3 expressions were measured by Western blotting. RESULTS: MTT results indicated that the vitality of ESCs in the active ingredients of PT groups at 48 h was increased compared with the control group and 2B had better effects than the others. FCM results indicated that 2B had the most significant anti-apoptotic effect compared with the control as well as S6, S8 and S9. Western blot results indicated that 2B, S6, S8 and S9 up-regulated the expression of Bcl-2 protein and down-regulated the expression of caspase-3 protein compared with the control. CONCLUSION: Ethyl acetate extract of Plastrum Testudinis inhibits epidermal stem cell apoptosis in serum-deprived culture by regulating the expressions of Bcl-2 and caspase-3 proteins and has a stronger anti-apoptotic effect than its constituents S6, S8 and S9.

[Effects and mechanism of Plastrum testudinis extracts on PC12 apoptosis].

OBJECTIVE: To observe the inhibitive effects of Plastrum testudinis Extracts (PTE) on 6-Hydroxydopamine (6-OHDA) induced PC12 cells apoptosis and explore its mechanism. METHODS: PC12 apoptosis model was established by serum starvation and damaged for 24 hours. The cells were randomly divided into four groups:control group, 6-OHDA group, PTE 3, 30 microg/mL group. Cell optical density was determined by MTT; Ratio of cell apoptosis was examined by Annexin V/PI double stain flow cytometry (FCM), and Western blot was applied to detect the BCL-X/L expression. RESULTS: MTT and FCM analysis demonstrated that PTE can elevate PC12 cells viability and reduce their apoptotic ratio in a dose dependent manner. Western blot showed that PTE promoted the expression of BCL-X/L. CONCLUSION: PTE can inhibit the apoptosis of PC12 induced by 6-OHDA in a dose dependent manner, and its mechanism maybe associated partially with up-regulating BCL-X/L signaling pathway.

BMP-Id pathway targeted by cholesterol myristate suppresses the apoptosis of PC12 cells.

Identifying small molecules that suppress apoptosis is promising for the therapy of brain diseases. We recently showed that autocrine bone morphogenetic protein (BMP) signaling involves the effects of cholesterol myristate present in traditional Chinese medicine on mesenchymal stem cells. The present study evaluated the effects of cholesterol myristate on the apoptosis and BMP signaling of PC12 cells. PC12 cells transfected by the inhibitor of differentiation (Id1) promoter reporter construct target gene of BMP4 signaling; cholesterol myristate increases the activity of Id1 promoter. However, structurally related steroids such as cholesterol, ß-sitosterol and cholesten-3-one, lack of the myristate, did not affect the activity of Id1 promoter, suggesting that myristate is essential for the effect of cholesterol myristate. These effects depend on BMP signaling. Apoptosis analysis indicated that cholesterol myristate inhibited the apoptosis of PC12 cells induced in serum-free condition. Cholesterol myristate significantly increases the expression of BMP4, BMPRIA, p-Smad1/5/8, Id1 and its antiapoptotic target gene Bcl-xL in PC12 cells treated in serum-free condition. Moreover, BMP antagonist reduced the anti-apoptotic effect of cholesterol myristate. Thus, this study is to provide evidence that BMP-Id pathway targeted by cholesterol myristate suppresses the apoptosis of PC12 cells. Our findings are therefore of considerable therapeutic significance and provide the potential of newly exploiting cholesterol myristate and clinically in brain disease therapies.

(+)-Cholesten-3-one induces differentiation of neural stem cells into dopaminergic neurons through BMP signaling.

To identify small molecules that induce dopaminergic neurons from neural stem cells (NSCs) is promising for therapy of Parkinson's disease. Here we report the results of analyzing structurally related steroids in traditional Chinese medicine to identify agents that enhance dopaminergic differentiation of NSCs. Using P19 cells transfected by tyrosine hydroxylase (TH) promoter reporter construct, (+)-Cholesten-3-one with carbonyl, but not cholesterol and cholesterol myristate can effectively promote the activity of TH promoter. This effect depends on bone morphogenetic protein (BMP) signaling. Phenotypic cellular analysis indicated that (+)-Cholesten-3-one induces differentiation of NSCs to dopaminergic neurons with increased expression of specific dopaminergic markers including TH, dopamine transporter, dopa decarboxylase and higher level of dopamine secretion. (+)-Cholesten-3-one significantly increases the expression of BMPR IB, but not BMPR IA or BMPR II; p-Smad1/5/8 positive nuclei and expression of p-Smad1/5/8 were detected in NSCs treated with (+)-Cholesten-3-one, indicating that (+)-Cholesten-3-one may activate the BMP signaling. Moreover, overexpression of BMP4 or inhibition of BMP affects the effect of (+)-Cholesten-3-one on the dopaminergic phenotype. These findings may contribute to efficient production of dopaminergic neurons from NSCs culture for many applications and raise interesting questions about the role of (+)-Cholesten-3-one in neurogenesis.

Hexadecanoic acid from Buzhong Yiqi decoction induced proliferation of bone marrow mesenchymal stem cells.

Buzhong Yiqi decoction (BYD) is a well-known ancient tonic prescription in traditional Chinese medicine (TCM). The purpose of this study is to identify active components of BYD involved in promoting proliferation of mesenchymal stem cells (MSCs) and to investigate its mechanism. BYD was extracted with petroleum ether, ethanol, and water. Evidence provided by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, bromodeoxyuridine, proliferation cell nuclear antigen immunoreactivity, cell cycle analysis, and gas chromatography-mass spectrometry indicated that hexadecanoic acid (HA) in BYD extracted with petroleum ether is the active compound responsible for increasing proliferation of MSCs. Western blot analysis show that HA significantly increase retinoic acid receptor (RAR) levels of MSCs, but not estrogen receptor, thyroid hormone receptor, vitamin D receptor, glucocorticoid receptor, and peroxisome proliferator-activated receptor. Reverse transcription-polymerase chain reaction revealed that HA significantly increased RAR mRNA levels. Furthermore, the mechanism of HA action depends on RAR pathway and up-regulates expression of mRNA for insulin-like growth factor-I, the target gene of RAR. Our findings have now allowed for a refinement in our understanding of TCM with respect to pharmacological regulation of stem cells and may be useful to stem cell biology and therapy.

Autocrine BMP4 signaling involves effect of cholesterol myristate on proliferation of mesenchymal stem cells.

We recently identified that cholesterol myristate in traditional Chinese medicine (TCM) is the active compound that increases proliferation of mesenchymal stem cell (MSCs). The present study is further to determine what signal pathway involves in effect of cholesterol myristate. Reverse transcription-PCR, Western blot and ELISA analysis show that cholesterol myristate increases the release of bone morphogenetic protein 4 (BMP4) from MSCs and the expression in the intracellular levels of BMP4 in a time- and dose dependent manner. However, structurally related steroids such as cholesterol and cholesten presented in TCM, both lack of the myristate, did not affect the secretion and expression of BMP4 on MSCs. These finds suggest that myristate is essential for the effects of cholesterol myristate. Furthermore, cholesterol myristate significantly increase BMPRIB levels of MSCs and the number of BMPRIB positive cells in a time- and dose dependent manner, but not BMPR IA or BMPR II. Our results indicate that action of cholesterol myristate may activate the BMP4-BMPRIB autocrine. Moreover, a blocking antibody against BMP4 or the BMP4 antagonist, noggin, partially reduced the effects of cholesterol myristate on MSCs proliferation. Thus, this study is to provide evidence that autocrine BMP4 signaling involves effect of cholesterol myristate on MSCs proliferation.

[Effects of Niupo Zhibao Pellet on high-mobility group box-1 protein expression in lung tissues of endotoxin shock rats].

OBJECTIVE: To observe the effects of Niupo Zhibao Pellet, a compound traditional Chinese herbal medicine, on high-mobility group box-1 protein (HMGB1) expression in lung tissues of rats with endotoxin shock. METHODS: Thirty SPF Sprague-Dawley rats were randomly divided into control group, lipopolysaccharide (LPS) group and Niupo Zhibao Pellet group. Rats in Niupo Zhibao Pellet group were consecutively administered 7 days with 3 mL (1 g/L) Niupo Zhibao Pellet saline suspension every day by intragastric administration. Endotoxin shock was induced in rats of the LPS and Niupo Zhibao Pellet groups by intravenous injection of LPS (1.5 mg/kg) and intraperitoneal injection of D-galactosamine (100 mg/kg). Expression of HMGB1 in lung tissues was measured by immunohistochemical method with diaminobenzidine (DAB) coloration, fluorescein isothiocyanate (FITC) labeling, and by Western blotting. RESULTS: Expression of HMGB1 in lung tissues in the LPS group was increased and that in Niupo Zhibao Pellet group was higher than that in the LPS group and the control group. HMGB1 was presented in the cytoplasm of positive cells in the LPS group, but in the nucleus of positive cells in the Niupo Zhibao Pellet group. However, HMGB1 was little expressed in the lung tissues of normal rats. CONCLUSION: Niupo Zhibao Pellet can increase HMGB1 expression and locate HMGB1 in the nucleus but not the cytoplasm, which may be one of its mechanisms in reducing endotoxin shock.

Characterization of chemical components in extracts from Si-wu decoction with proliferation-promoting effects on rat mesenchymal stem cells.

Si-wu decoction (SDE), a classic prescription in Traditional Chinese Medicine, has been used for the treatment of a variety of anaemia in China for centuries. In order to explore the scientific basis of the formula, we investigated the relationship between its chemical components and proliferation-promoting effects on rat marrow-derived mesenchymal stem cells (MSCs). Twenty (F-1-F-20) components were obtained and their proliferation-promoting effects on MSCs were investigated. The results showed that F-4, F-7, F-10, and F-11 stimulated the proliferation of the MSCs. The chemical components with proliferation-promoting effects on the MSCs were further identified by GC-MS, HPLC, LC-MS, and other spectra. Ligustilide (F-4) isolated from SDE showed the best proliferation-promoting effect. Palmitic acid methyl ester and stearic acid ethyl identified from F-7 and F-10 by HPLC were also confirmed to be responsible for stimulating MSC proliferation. A novel compound, 6,7-dihydroxy-3-(-prop-1-enyl)isobenzofuran-1(3H)-one, was found in the SDE for the first time by LC-MS(n), whose structure was similar to ligustilide.