Modified Xiaochaihu Decoction () Promotes Collagen Degradation and Inhibits Pancreatic Fibrosis in Chronic Pancreatitis Rats.

OBJECTIVE: To investigate the effect of Modified Xiaochaihu Decoction (MXD, ) on collagen degradation in rats with chronic pancreatitis (CP). METHODS: Rats were injected dibutyltin dichloride (DBTC, 7 mg/kg of body weight) into the right caudal vein to induce CP model. Thirty heallhy male Wistar rats were randomly divided into three groups by a random number table: the control, the model and the treatment groups. Rats of treatment group were administered MXD (10 g/kg of body weight) orally once daily starting from the day post-model establishment. Pancreatic tissues were harvested after 28-day feeding and fibrosis was evaluated by picro-sirius red staining. The contents of collagen type I and III were detected using enzymelinked immunosorbent assay (ELISA), the expression of matrix metalloproteinase 13 (MMP13) and tissue inhibitor of metalloproteinase 1 (TIMP1) was analyzed by Western blot and real-time polymerase chain reaction (PCR). RESULTS: The fibrosis scoring of pancreatic tissues, the concentrations of collagen type I and III, the expression levels of MMP13 and TIMP1 proteins and mRNA in the model group were all increased compared with the control group (P<0.05). After treatment with MXD, the fibrosis scoring of pancreatic tissues, the concentrations of collagen type I and III, the expression levels of MMP13 proteins and mRNA in the teatment group were all decreased compared with the model group (P<0.05), but there were no significant differences in the expression levels of TIMP1 proteins and mRNA (P>0.05). CONCLUSIONS: MXD could promote collagen degradation and reverse pancreatic fibrosis in CP rats via a mechanism involve up-regulation of MMP13 expression.

Modified Da-chai-hu Decoction regulates the expression of occludin and NF-κB to alleviate organ injury in severe acute pancreatitis rats.

Modified Da-chai-hu Decoction (MDD), a traditional Chinese medicinal formulation, which was empirically generated from Da-chai-hu decoction, has been utilized to treat severe acute pancreatitis (SAP) for decades. The aim of the present study was to explore its potential organprotective mechanism in SAP. In the present study, rat SAP model was induced by retrograde injection of 3.5% sodium taurocholate into the biliopancreatic duct, MDD (23.35 g/kg body weight, twelve times the clinical dose) were orally given at 2 h before and 10 h after injection. At 12 h after model induction, blood was taken from vena cava for analysis of amylase, diamine oxidase (DAO), pulmonary surfactant protein-A (SP-A), and C-reactive protein (CRP). Histopathological change of pancreas, ileum and lung was assayed by H&E staining, myeloperoxidase (MPO) activity were determinated using colorimetric assay, and the expressions of occludin and nuclear factor-κB (NF-κB) were detected by real-time RT-PCR and western blot, respectively. In addition, the tissue concentrations of tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), and monocyte chemoattractant protein-1 (MCP-1) were measured by enzyme-linked immunosorbent assay (ELISA). The results showed that in SAP rats, MDD significantly alleviated histopathological damage, depressed the MPO activity and the concentrations of TNF-α, IL-1ß, and MCP-1 of pancreas, ileum and lung, and reduced the serum levels of amylase [(3283.4 ± 585.5) U·L-1vs (5626.4 ± 795.1)U·L-1], DAO [(1100.1 ± 334.3) U·L-1vs (1666.4 ± 525.3) U·L-1] and CRP [(7.6 ± 1.2) µg·mL-1vs (17.8 ± 3.8) µg·mL-1]. However, the serum SP-A concentration [(106.1 ± 16.6) pg·mL-1vs (90.1 ± 14.9) pg·mL-1] was elevated when treated SAP rats with MDD. Furthermore, MDD increased the occludin expression and reduced the NF-κB expression in pancreas, ileum and lung of SAP rats. Our findings suggested that MDD administration was an effective therapeutic approach for SAP treatment. It could up-regulate occludin expression to protect intercellular tight junction and down-regulate NF-κB expression to inhibit inflammatory reaction of pancreas, ileum and lung.

Saikosaponin d ameliorates pancreatic fibrosis by inhibiting autophagy of pancreatic stellate cells via PI3K/Akt/mTOR pathway.

Chronic pancreatitis is characterized by pancreatic fibrosis, associated with excessive activation of pancreatic stellate cells (PSCs) and increased expression of transforming growth factor-ß1 (TGF-ß1). Recently, our studies have shown that autophagy inhibitor could inhibit PSCs activation and reduce collagen secretion. Saikosaponin d (SSd), the major active component of bupleurum falcatum (a medicinal plant), has anti-fibrosis effects in liver. However, it is unclear whether SSd has a role in pancreatic fibrosis. This study aimed to investigate the effect of SSd on the autophagy and activation of PSCs in vivo and in vitro. In vivo, a rat chronic pancreatitis model was induced by intravenous injection of dibutyltin dichloride. SSd was administered at a dose of 2.0 mg/kg body weight per day by gavage. After 4 weeks, the pancreas was collected for histological and molecular analysis. In vitro, PSCs were isolated and cultured for treatment with different dosages of SSd. The results showed that SSd inhibited PSCs autophagy and activation while also reducing extracellular matrix (ECM) formation and pancreatic damage. SSd inhibited autophagy through activating the PI3K/Akt/mTOR pathway. SSd also promoted degradation of ECM with an increasing ratio of MMPs/TIMPs and suppressed the TGF-ß1/Smads pathway. From these results, we concluded that SSd prevents pancreatic fibrosis by reducing autophagy of PSCs through PI3K/Akt/mTOR pathway, which has crosstalk with the TGF-ß1/Smads pathway.

Modified Xiaochaihu Decoction () prevents the progression of chronic pancreatitis in rats possibly by inhibiting transforming growth factor-ß1/Sma- and mad-related proteins signaling pathway.

OBJECTIVE: To investigate the effect of modified Xiaochaihu Decoction (, MXD) on transforming growth factor-ß1/Sma- and Mad-related proteins (TGF-ß1/Smads) signaling pathway in rats with chronic pancreatitis (CP) induced by dibutyltin dichloride. METHODS: Thirty healthy male Wistar rats were randomly divided into the normal control group, CP group and CP+MXD-treated group. CP was induced by injection of dibutyltin dichloride (DBTC, 7 mg/kg of body weight) into the right caudal vein, and the control rats were treated with vehicle. MXD was given daily by gavage at a dose of 10 g/kg of body weight, starting from the day after CP induction. After 28-day treatment, the n-benzoyl-tyrosyl para-aminobenzoic acid (NBT-PABA) test was carried out to evaluate exocrine pancreatic function. Then, rats were sacrificed, and pancreatic tissues were harvested for histological evaluation. In addition, the mRNA expression of TGF-ß1, TGF-ß1 type II receptor (TGFßRII), Smad3 and Smad7 was determined in pancreatic tissues by using real-time polymerase chain reaction. RESULTS: Treatment of CP with MXD improved the PABA recovery, decreased the histological lesion, and reduced the mRNA expression of TGF-ß1, TGFßRII and Smad3 (P<0.05). However, MXD had no effect on Smad7 mRNA level. CONCLUSIONS: MXD could protect the pancreas against chronic injury and improve pancreatic exocrine function in DBTC induced rat CP model. Its mechanism may involve inhibition of the TGF-ß1/Smads signaling pathway.

[Relationship between immune imbalance and sthenia-asthenia syndromes in patients suffering from severe intra-abdominal infection].

OBJECTIVE: To explore the characteristics of immune imbalance in patients with multiple organ dysfunction syndrome (MODS) induced by severe intra-abdominal infection and its relationship with changing of TCM sthenia-asthenia syndrome. METHODS: Forty-six patients with MODS induced by severe intra-abdominal infection and treated with etiological and syndrome differentiation of integrative medicine were observed in succession. Patients' peripheral blood levels of interleukin-6/interleukin-10 ratio (IL-6/IL-10), human leukocyte antigen DR site (HLA-DR), helper T lymphocyte1/2 ratio (Th1/Th2), and the regulatory T lymphocyte (Treg) were measured on the 1st, 3rd and 7th day of the research respectively. And the distribution laws of TCM syndrome types, sthenia (S), asthenia (A), and mingled sthenia/asthenia (M), in patients were observed as well. RESULTS: IL-6/IL-10 ratio at all the testing time points showed insignificant difference in patients of types S and M, while in those of type A, it was more lowered on the 7th day than that on the 1st day. HLA-DR lowered to <30% on the 7th day in all patients of type A and showed significant difference to that on the 1st day (P <0.05), while HLA-DR <30% was not found in all patients of types S and M. Th1/Th2 ratio in patients of types S and A was insignificant different at the foremost 3 days, but lowered significantly on the 7th day, while in patients of type M, it was unchanged in all the 7 days of observation. Treg level was unchanged in the foremost 3 days in patients of types S and M, while in those of type A, it raised on the 3rd day, but no raising was found in the subsequent 4 days. Comparisons of various indexes detected at corresponding time points respectively among patients with various syndrome types showed that, for levels of IL-6/IL-8, HLA-DR, and Th1/Th2, the sequence was S>M>A; and for Treg, it was A>M>S. CONCLUSION: In the pathological process of MODS induced by severe intra-abdominal infection, the index IL-6/IL-10, reflecting the balance of the pro-/anti-inflammatory cytokines and the indexes HLA-DR, Th1/Th2 and Treg reflecting the immune function, all can exactly reflect the TCM asthenia-sthenia syndrome types. The sequence in patients of various syndrome types for levels of IL-6/IL-10, HLA-DR and Th1/Th2, is S> M>A, but for Treg it is the inverse, as A>M>S.

[Effect of clearing heat and removing dampness method on formation of pigment gallstones in rabbits].

OBJECTIVE: To observe dynamically the effect of drugs for clearing heat and removing dampness (CHRD) on biliary components in rabbits with pigment gallstones (PGS). METHODS: Forty rabbits were established into PGS model and randomly divided into 3 groups, the bacterial infection group, the CHRD low-dose group and the CHRD high-dose group. Besides, a normal group was set up with healthy rabbits for control. Changes of total bilirubin (TB), unconjugated bilirubin (UCB), total bile acid (TBA), Ca2+, bacterial and endogenous beta-glucuronidase (beta-Gase) in bile were observed. RESULTS: CHRD drugs significantly decreased the contents of UCB, Ca2+, bacterial and endogenous beta-Gase (P < 0.05), and increased TBA in bile (P < 0.05). CONCLUSION: CHRD drugs have good effect in reducing the lithogenesis of the pigment gallstones.

Effects of choleretics on bile compositions drained from patients with pigment gallstone.

OBJECTIVE: To provide evidence for three-level prevention of cholelithiasis by means of observing the effects of some choleretics on bile compositions drained from patients with pigment gallstone. METHODS: Twenty-seven patients suffering from primary pigment gallstones and having received treatment of choledochostomies plus T-tube or endoscopic nasal bile drainage (ENBD) were divided equally into three groups, and administered respectively with Lidanling (the LDL group), ursodesoxycholic acid (the UDA group) and combination of LDL and UDA (the LDL + UDA group) through oral intake (7 patients in each group). Besides, 6 post-operational patients got no treatment with any drug were allocated in the control group. Bile of all the patients was collected before treatment and on the 1, 3, 5, 7 th day after the treatment started to detect levels of total bile acid (TBA), glycocholic acid (GCA), taurocholic acid (TCA), glycocholic cheno-desoxycholic acid (GCDCA), total bilirubin (TBIL), uncombined bilirubin (UCB), concentration of calcium ion (Ca(2+)) as well as the bacterio-genetic and endogenous beta-glucuronidase activity for comparing. RESULTS: Levels of TBA, GCA, TCA and GCDCA got gradually increased in the UDA group and the LDL + UDA group after treatment (P < 0.05), while those in the LDL group remained unchanged, showing an insignificant difference as compared with those in the control group. In the LDL group and the LDL + UDA group, TBIL gradually increased while UCB gradually decreased in the course of treatment (P < 0.05). Moreover, levels of Ca(2+) and endogenous beta-glucuronidase activity got significantly lowered (P < 0.05). CONCLUSION: Combined use of LDL and UDA could elevate levels of TBA, GCA, TCA, GCDCA, enhance the excretion of TBIL in patients with pigment gallstone after bile drainage, lower levels of UCB and Ca(2+) and the activity of endogenous beta-glucuronidase in the bile, so as to reduce the possibility of stone formation of bile, and therefore, it could be used to prevent the production of pigment gallstone, especially to prevent post-operative recurrence of stones.

[Monocyte response and regulatory effect of emodin and shenmai injection on it in patients with severe sepsis].

OBJECTIVE: To investigate the impact of monocyte releasing tumor necrosis factor-alpha (TNF-alpha) and interleukin-10 (IL-10) stimulated by lipopolysaccharide (LPS) in severe sepsis (SS) patients, and the regulatory effect of emodin and Shenmai Injection (SMI) on monocyte response. METHODS: ocyte (PBMC) sampled from SS patients due to severe abdominal inflammation and healthy controls, PBMC were incubated with or without LPS, respectively. PBMC media pretreated with emodin and SMI, and then the levels of cytokine factors including TNF-alpha and IL-10 in supernatants were determined after stimulated with or without LPS in the two groups. Normal PBMC stimulated with LPS based on incubated with either normal serum or SS serum, and the levels of TNF-alpha and IL-10 in supernatant of normal PBMC and SS serum dealing with emodin or SMI after cultured and stimulated with LPS were determined, respectively. RESULTS: The levels of TNF-alpha and IL-10 were significantly higher in SS patients than those in the healthy adults. The contents of TNF-alpha and IL-10 increased in response to LPS in PBMC of healthy adults, and the excretion of TNF-alpha was significantly attenuated whereas IL-10 significantly increased in septic PBMC than basal content. Both of the two traditional Chinese medicines had significantly effect in stimulating PBMC secretion in healthy adults and SS patients. In normal PBMC, emodin attenuated TNF-alpha and IL-10 release in response to LPS, and SMI significantly inhibited TNF-alpha release. As to septic PBMC, emodin significantly stimulated TNF-alpha and IL-10 release and SMI significantly increased the concentration of TNF-alpha in the SS patients. Incubation of normal PBMC with septic serum attenuated TNF-alpha production, whereas increased IL-10 release. Emodin could significantly decrease the level of IL-10, and SMI recovered TNF-alphareleasing and had no effect on IL-10. CONCLUSION: The level of TNF-alpha significantly decreased accompanied with an anti-inflammatory cytokine IL-10 significantly increased of PBMC in SS patients. Monocyte exhibits the different response of inflammatory or anti-inflammatory factor which may be one of the reasons of imbalance of immune function in SS patients. Both of emodin and SMI may have regulatory effect on excretion of PBMC in SS patients, but they play a role in different ways.

Change of TH1/TH2 cytokine equilibrium in rats with severe sepsis and therapeutic effect of recombinant interleukin-12 and Shenmai injection.

OBJECTIVE: To evaluate the dynamic change of Th1/Th2 cytokines in serum, peritoneal lavage fluid (PLF) and splenic macrophages (SM) in rats with severe peritonitis, and to observe the therapeutic effects of recombinant interleukin-12 (rIL-2) and Shenmai injection (SMI), a Chinese medicinal preparation. METHODS: Severe peritonitis (SP) model was induced by intraperitoneal injection of E. coli and B. frag, and mild peritonitis (MP) model was induced by cecal ligation and punching. Then the following experiments were done: (1) Survival rates of animals after every 6 hrs in the 72 hrs after modeling were recorded, serum and PLF levels of cytokines, including tumor necrosis factor-alpha (TNF-alpha), interferon-gamma (IFN-gamma), and interleukin-10 (IL-10), 6 hrs, 12 hrs, 24 hrs and 48 hrs after modeling were measured. (2) Model rats were treated with rIL-12 or SMI, the survival rate was recorded and serum levels of TNF-alpha, INF-gamma, and IL-10 before and after treatment were measured, and (3) amount of these cytokines produced by SM were determined 6 hrs, 12 hrs and 24 hrs after treatment. The survival rates and levels of cytokines were then compared between the groups (model group treated with rIL-12 or SMI, untreated model group, and blank group). RESULTS: Serum and PLF levels of IFN-gamma, TNF-alpha at all the time points in SP rats were significantly lower than those in MP rats while those of IL-10 6 hrs and 12 hrs after modeling were significantly higher in the former than that in the latter (P < 0.05). IFN-gamma secretion of SM in SP rats was significantly higher than that in MP rats 6 hrs after modeling (P < 0.05). Administration of rlL-12 or SMI given before modeling could improve the survival rate of the model rats (P < 0.05) and cause significant increase of the serum level and SM secretion of IFN-gamma. CONCLUSION: Imbalance in promoting/antagonizing inflammatory cytokines and Th2 response dominance appear in SP rats early at the initiating stage, and SM secretion of inflammation promoting factor also reduces. Administration in time of rIL-12 and SMI, may increase the survival rate, and its mechanism may be related with their immuno-stimulating action.

Anti-inflammatory effects of an herbal medicine (Xuan-Ju agent) on carrageenan- and adjuvant-induced paw edema in rats.

Xuan-Ju agent is an herbal formula containing aqueous extract of Formica fusca, Herba epimedii, Fructus cnidii, and Fructus lycii, all of which are reputed for their beneficial effects in the treatment of the immunodeficient diseases such as rheumatoid arthritis. We performed a study on the anti-inflammatory effects of this agent using carrageenan- and adjuvant-induced paw edema in rats. Xuan-Ju agent showed a marked inhibitory effect on edema in two models of inflammation in rats, at the dose of 0.20, 0.40 and 0.80g/kg. Based on this study, Xuan-Ju agent is considered to be a potentially useful drug suitable for further evaluation for rheumatoid arthritis.