Ethanol extract of Herpetospermum caudigerum Wall ameliorates psoriasis-like skin inflammation and promotes degradation of keratinocyte-derived ICAM-1 and CXCL9.

OBJECTIVE: To explore whether the ethanol extract of Herpetospermum caudigerum Wall (EHC), a Xizang medicinal plant traditionally used for treating liver diseases, can improve imiquimod-induced psoriasis-like skin inflammation. METHODS: Immunohistochemistry and immunofluorescence staining were used to determine the effects of topical EHC use in vivo on the skin pathology of imiquimod-induced psoriasis in mice. The protein levels of interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α), and interleukin-17A (IL-17A) in mouse skin samples were examined using immunohistochemical staining. In vitro, IFN-γ-induced HaCaT cells with or without EHC treatment were used to evaluate the expression of keratinocyte-derived intercellular cell adhesion molecule-1 (ICAM-1) and chemokine CXC ligand 9 (CXCL9) using Western blotting and reverse transcription-quantitative polymerase chain reaction. The protein synthesis inhibitor cycloheximide and proteasome inhibitor MG132 were utilized to validate the EHC-mediated mechanism underlying degradation of ICAM-1 and CXCL9. RESULTS: EHC improved inflammation in the imiquimod-induced psoriasis mouse model and reduced the levels of IFN-γ, TNF-α, and IL-17A in psoriatic lesions. Treatment with EHC also suppressed ICAM-1 and CXCL9 in epidermal keratinocytes. Further mechanistic studies revealed that EHC suppressed keratinocyte-derived ICAM-1 and CXCL9 by promoting ubiquitin-proteasome-mediated protein degradation rather than transcriptional repression. Seven primary compounds including ehletianol C, dehydrodiconiferyl alcohol, herpetrione, herpetin, herpetotriol, herpetetrone and herpetetrol were identified from the EHC using ultra-performance liquid chromatography-quadrupole-time of flight-mass spectrometry. CONCLUSION: Topical application of EHC ameliorates psoriasis-like skin symptoms and improves the inflammation at the lesion sites. Please cite this article as: Zhong Y, Zhang BW, Li JT, Zeng X, Pei JX, Zhang YM, Yang YX, Li FL, Deng Y, Zhao Q. Ethanol extract of Herpetospermum caudigerum Wall ameliorates psoriasis-like skin inflammation and promotes degradation of keratinocyte-derived ICAM-1 and CXCL9. J Integr Med. 2023; 21(6): 584-592.

Resting-state fMRI reveals changes within the anxiety and social avoidance circuitry of the brain in mice with psoriasis-like skin lesions.

Psoriasis is an autoimmune skin disease that often co-occurs with psychological morbidities such as anxiety and depression, and psychosocial issues also lead psoriasis patients to avoid other people. However, the precise mechanism underlying the comorbidity of psoriasis and anxiety is unknown. Also, whether the social avoidance phenomenon seen in human patients also exists in psoriasis-like animal models remains unknown. In the present study, anxiety-like behaviours and social avoidance-like behaviours were observed in an imiquimod-induced psoriasis-like C57-BL6 mouse model along with typical psoriasis-like dermatitis and itch-like behaviours. The 11.7T resting-state functional magnetic resonance imaging showed differences in brain regions between the model and control group, and voxel-based morphometry showed that the grey matter volume changed in the basal forebrain region, anterior commissure intrabulbar and striatum in the psoriasis-like mice. Seed-based resting state functional connectivity analysis revealed connectivity changes in the amygdala, periaqueductal gray, raphe nuclei and lateral septum. We conclude that the imiquimod-induced psoriasis-like C57-BL6 mouse model is well suited for mechanistic studies and for performing preclinical therapeutic trials for treating anxiety and pathological social avoidance in psoriasis patients.

Research on the wound healing effect of Shengji Huayu Formula ethanol extract-derived fractions in streptozotocin-induced diabetic ulcer rats.

BACKGROUND: Diabetic ulcer is a common complication of diabetes. It is characterized by a long-term disease course and high recurrence rate. Shengji Huayu Formula (SHF) is an effective formula for treating diabetic ulcers. However, the specific effective parts of SHF remain unclear. Clarifying the active polar site of SHF would be helpful to refine research on the components in SHF that promote wound healing. This research aims to focus on evaluating the activity of polar fractions. METHODS: A diabetic rat model was established by intraperitoneally injecting streptozotocin (STZ) and was adopted to confirm the therapeutic effect of SHF. Four different polarity parts were extracted from SHF and prepared into a cream to evaluate the activity. High-performance liquid chromatography (HPLC) was used to detect chemical constituents in chloroform extracts. RESULTS: It was discovered that dracorhodin, aloe-emodin, rhein, imperatorin, emodin, isoimperatorin, chrysophanol, physcion, and tanshinone IIA were the main components of the chloroform extract from SHF. The results revealed that chloroform extract could effectively accelerate diabetic wound healing by promoting collagen regeneration and epidermal repair. Chloroform extract of SHF could stimulate the generation of vascular endothelial growth factor (VEGF). The results are also indicated that the effective active fraction was the chloroform part, and the method of detecting the main chemical constituents in the active part was successfully established. CONCLUSION: SHF could improve diabetic ulcers by promoting granulation tissue synthesis. In this study, four polar parts (petroleum ether, chloroform, ethylacetate, n-butanol) were extracted from a 95% ethanol extract. In contrast, chloroform polar parts showed a higher wound closure rate, stimulated more collagen regeneration and promoted more production of vascular endothelial cells. In conclusion, the chloroform extract of SHF was the effective polar part in ameliorating diabetic wound healing.

Therapeutic effects of the extract of Sancao Formula, a Chinese herbal compound, on imiquimod-induced psoriasis via cysteine-rich protein 61.

OBJECTIVE: Psoriasis is a common chronic inflammatory skin disease that is prone to recurrence, and the proinflammatory factor, cysteine-rich protein 61 (Cyr61), is important in its pathophysiology. Long-term clinical practice has shown that Sancao Formula (SC), a Chinese herbal compound, is effective in the treatment of psoriasis, but the precise mechanism remains unknown. In this study, we investigate the mechanism by which SC extract alleviates imiquimod (IMQ)-induced psoriasis. METHODS: The expression of Cyr61 in psoriatic lesions and normal healthy skin was detected using immunohistochemical analysis to investigate the biological role of Cyr61 in models of psoriatic inflammation. A psoriatic mouse model was established by topical application of IMQ, and the effect of topical application of SC extract was evaluated using the psoriasis area and severity index (PASI) score, hematoxylin-eosin staining, and histopathological features of the skin. Next, a HaCaT cell inflammation model was established using interferon-γ (IFN-γ), and the effect of SC extract on the mRNA and protein levels of Cyr61 and intercellular cell adhesion molecule-1 (ICAM-1) was confirmed using Western blot and quantitative real-time polymerase chain reaction analyses. RESULTS: Immunohistochemical staining showed that the expression of Cyr61 in psoriatic lesions was higher than that in normal skin samples (78.26% vs 41.18%, P < 0.05), and the number of Cyr61-positive cells in psoriatic lesions was also significantly higher than in normal skin (18.66 ± 2.51 vs 4.33 ± 1.52, P < 0.05). Treatment in mice with IMQ-induced psoriasis showed that SC extract could significantly improve the inflammatory phenotype, PASI score (10.875 ± 0.744 vs 3.875 ± 0.582, P < 0.05), and pathological features compared with those in IMQ model group; SC treatment was also associated with decreased levels of Cyr61 and ICAM-1. In the IFN-γ-induced inflammatory cell model, the mRNA and protein levels of Cyr61 and ICAM-1 were upregulated, while the SC extract downregulated the levels of Cyr61 and ICAM-1. CONCLUSION: The results provide a theoretical basis for the involvement of Cyr61 in the pathogenesis of psoriasis, and suggest that SC should be used to target Cyr61 for the prevention of psoriasis recurrence.

Application of a Cloud Model-Set Pair Analysis in Efficacy Assessment for Diabetic Ulcers.

Because treatment of diabetic ulcers includes various uncertainties, efficacy assessments are needed and significant. In previous studies, set pair analysis (SPA) has been applied to the efficacy assessments of traditional Chinese medicine (TCM) that pick out uncertainties related to the development and prognosis of disease. Optimized clinical protocols of SPA improve clinical efficacy. In the article, cloud model (CM) is employed to improve SPA, and a novel efficacy assessment method for a treatment of diabetic ulcers is proposed based on the cloud model-set pair analysis (CM-SPA). It is recommended to replace connection degree (CD) with cloud connection degree (CCD) that the efficacy assessment results are shown as normal clouds. Then, three diabetic ulcers patients treated with TCM made importance assessment by both CM-SPA and AHP based SPA. The comparison of assessment results shows that the CM-SPA is efficacious for the efficacy assessment of a treatment for diabetic ulcers and the results will be more scientific and accurate via CM-SPA.

Acute and chronic toxicity of a polyherbal preparation - Jueyin granules.

BACKGROUND: The potential toxicity of Chinese herbal medicine has attracted more attention in recent years. Jueyin granules (JYG), a polyherbal formula, have been proven to be an effective agent for treating psoriasis in both animal models and clinical research. However, little is known about the possible acute and chronic toxicity of JYG. The objective of this study was to investigate the safety of JYG in ICR mice and Wistar rats. METHODS: To examine the acute toxicity of JYG, ICR mice were randomly divided into an experimental group and a control group, each comprising 20 mice (10 male and 10 female). The experimental group was fed JYG solution at a dose of 21.5 g/kg, equivalent to 143 times the clinical human dosage, for 14 days, whereas control animals were fed distilled water. In the chronic toxicity test, Wistar rats were divided into four groups, each comprising 40 rats (20 male and 20 female). For 6 months, the experimental animals were given JYG at a dose of 7.5, 3.75 and 1.875 g/kg, whereas control animals were given distilled water. The animals' body weight, food and water consumptions were monitored weekly. In addition, their biochemical and hematological parameters, histopathology, and body and organ weights were all measured at specific observation time points. RESULTS: According to the results of the acute toxicity test, no mortality was found and no abnormal pathological changes in major organs were observed in mice treated with JYG. In the chronic toxicity test, JYG did not cause significant abnormalities in the physiological parameters or pathological changes in the major organs of the rats. CONCLUSION: The results indicated that JYG at the given doses did not induce any harmful effects in animals. Thus, it is reasonable to conclude that JYG is safe at the studied dosage levels and causes no acute or chronic toxicity in animal models.

Sheng-ji Hua-yu formula promotes diabetic wound healing of re-epithelization via Activin/Follistatin regulation.

BACKGROUND: Sheng-ji Hua-yu(SJHY) formula is one of the most useful Traditional Chinese medicine (TCM) in the treatment of the delayed diabetic wound. However, elucidating the related molecular biological mechanism of how the SJHY Formula affects excessive inflammation in the process of re-epithelialization of diabetic wound healing is a task urgently needed to be fulfilled. The objectives of this study is to evaluate the effect of antagonisic expression of pro-/anti-inflammatory factors on transforming growth factor-ß(TGF-ß) superfamily (activin and follistatin) in the process of re-epithelialization of diabetic wound healing in vivo, and to characterize the involvement of the activin/follistatin protein expression regulation, phospho-Smad (pSmad2), and Nuclear factor kappa B p50 (NF-kB) p50 in the diabetic wound healing effects of SJHY formula. METHODS: SJHY Formula was prepared by pharmaceutical preparation room of Yueyang Hospital of Integrated Traditional Chinese and Western Medicine. Diabetic wound healing activity was evaluated by circular excision wound models. Wound healing activity was examined by macroscopic evaluation. Activin/follistatin expression regulation, protein expression of pSmad2 and NF-kB p50 in skin tissue of wounds were analyzed by Real Time PCR, Western blot, immunohistochemistry and hematoxylin and eosin (H&E) staining. RESULTS: Macroscopic evaluation analysis showed that wound healing of diabetic mice was delayed, and SJHY Formula accelerated wound healing time of diabetic mice. Real Time PCR analysis showed higher mRNA expression of activin/follistatin in diabetic delayed wound versus the wound in normal mice. Western Blot immunoassay analysis showed reduction of activin/follistatin proteins levels by SJHY Formula treatment 15 days after injury. Immunohistochemistry investigated the reduction of pSmad2 and NF-kB p50 nuclear staining in the epidermis of diabetic SJHY versus diabetic control mice on day 15 after wounding. H&E staining revealed that SJHY Formula accelerated re-epithelialization of diabetic wound healing. CONCLUSION: The present study found that diabetic delayed wound healing time is closely related to the high expression level of activin/follistatin, which leads to excessive inflammation in the process of re-epithelization. SJHY Formula accelerates re-epithelialization and healing time of diabetic wounds through decreasing the high expression of activin/follistatin.

Immune Signatures in Patients with Psoriasis Vulgaris of Blood-Heat Syndrome: A Systematic Review and Meta-Analysis.

Objective. To determine whether immunological serum markers IFN-γ, IL-4, IL-17, IL-23, IL-6, TNF-α, and IL-10 are elevated or decreased in patients compared with healthy controls. Methods. A complete search of the literature on this topic within the past 30 years was conducted across seven databases. Seventeen studies including 768 individuals were identified. Differences in serum marker levels between subjects and controls were pooled as MDs using the random-effects model. Results. The pooled MDs were higher in patients than in healthy controls for IFN-γ (MD 24.9, 95% CI 12.36-37.43), IL-17 (MD 28.92, 95% CI 17.44-40.40), IL-23 (MD 310.60, 95% CI 4.96-616.24), and TNF-α (MD 19.84, 95% CI 13.80-25.87). Pooled IL-4 (MD -13.5, 95% CI -17.74--9.26) and IL-10 (MD -10.33, 95% CI -12.03--8.63) levels were lower in patients. Conclusion. The pooled analyses suggest that levels of IFN-γ, IL-17, IL-23, and TNF-α are significantly elevated and that levels of IL-4 and IL-10 are significantly decreased in sera of patients with psoriasis vulgaris of blood-heat syndrome. Measuring progression of blood-heat syndrome of psoriasis vulgaris will require additional high-quality data, with a low risk of bias and adequate sample sizes, before and after antipsoriatic therapy.

External Application of Traditional Chinese Medicine for Venous Ulcers: A Systematic Review and Meta-Analysis.

Objective. To evaluate the effectiveness of external application of traditional Chinese medicine (EA-TCM) on venous ulcers. Methods. Seven databases were searched until April 2015 for randomized controlled trials (RCTs) of EA-TCM for venous ulcers. Risk of bias was assessed using Cochrane Handbook guidelines. Study outcomes were presented as risk ratios (RRs) for dichotomous data or mean differences (MDs) for continuous data. Results. Sixteen of 193 potentially relevant trials met the inclusion criteria; however, their methodological qualities were low. Comparison of the same intervention strategies revealed significant differences in total effectiveness rates between EA-TCM and conventional therapy groups (RR = 1.22, 95% confidence interval [CI] = 1.16-1.29, and P < 0.00001). Compared to conventional therapy, EA-TCM combined with conventional therapy had a superior total effectiveness rate (RR = 1.11, 95% CI = 1.04-1.19, and P = 0.003). There were no significant differences in recurrence rates during followup and final pain measurements between the experimental and those in the control groups (RR = 0.86, 95% CI = 0.31-2.39, and P = 0.85; MD -0.75, 95% CI = -2.15-0.65, and P = 0.29). Conclusion. The evidence that EA-TCM is an effective treatment for venous ulcers is encouraging, but not conclusive due to the low methodological quality of the RCTs. Therefore, more high-quality RCTs with larger sample sizes are required.

[Relation between Chinese Medical Constitutions of Female Patients with Late-onset Acne: an Epidemiological Investigation].

OBJECTIVE: To explore the correlation between syndrome types of late-onset acne female patients and constitutions of Chinese medicine (CM). METHODS: A questionnaire was performed in 365 late-onset acne female patients and 135 healthy subjects (as the control) using Professor WANG Qi's. methods and Standards for Chinese Medical Constitutions Classification. RESULTS: Their CM constitutions were sequenced as damp-heat constitution, yin-deficiency constitution, balanced constitution, yang-deficiency constitution, blood-stasis constitution, qi-stagnation constitution, qi-deficiency constitution, phlegm-damp constitution, inherited special constitution, with statistical difference when compared with those of the control group ( χ2 = 85.206, P < 0.01). In the 365 female late-onset acne patients, 114 (31.23%) were with Chongren imbalance syndrome, 108 (29.59%) were with blood stasis or coagulated phlegm syndrome, 83 (22.74%) were with dampness heat syndrome, and 60 (16.44%) were with wind heat syndrome. There was statistical difference in CM constitution distributions among different CM syndrome types (χ2 = 105.671, P < 0.01). The distribution of CM medical constitutions was different between the two groups. Biased constitutions were often seen in the patient group, while balanced constitution was often seen in the control group. Binary Logistic regression analysis indicated that influencing factors covered sweet food, light diet, roasted food, coffee, stress, work pressure, and family pressure. Of them light diet was one protective factor, while the rest were adverse factors. CONCLUSION: The etiology and syndrome types of female late-onset acne female patients were associated with CM constitution.

Expression of T-helper 17 cells and signal transducers in patients with psoriasis vulgaris of blood-heat syndrome and blood-stasis syndrome.

OBJECTIVE: To investigate the levels of cytokines related to T-helper (Th) 17 cells in serum and signal transducers in the psoriatic lesions of patients with psoriasis vulgaris of blood-heat syndrome (BHS) and blood-stasis syndrome (BSS). METHODS: Sixty patients with psoriasis vulgaris were divided into the BHS and BSS groups according to the syndrome differentiation of Chinese medicine (CM). Ten healthy subjects were considered as the control group. Cytokine levels of interleukin (IL)-17, IL-23 and IL-6 in serum were determined by enzyme-linked immunosorbent assay. Expression levels of signal transducer and activator of transcription 3 (STAT3), p38-mitogen-activated protein kinase (MAPK) and STAT6 in the psoriatic lesions were determined using immunohistochemistry (IHC), Western blot, and real-time quantitative reverse transcription polymerase chain reaction, respectively. RESULTS: Production of IL-17, IL-23 and IL-6 in the BHS group and BSS group were significantly increased compared with those in the control group (P<0.05). Levels of IL-17 and IL-23 in the BHS group were higher than those in the BSS group (P<0.05). Compared with the control group, IHC positive expressions and protein expressions of STAT3 and p38-MAPK, and the STAT3 mRNA expressions in the BHS and BSS groups were significantly higher (P<0.05 or P<0.01). The protein expression of STAT3 in the BHS group was significantly higher than that in the BSS group (P<0.05). CONCLUSIONS: Cytokines in serum and signal transducers in the psoriatic lesions alter with various CM syndromes of psoriasis. The results provide scientific basis for the treatment based on syndrome differentiation of CM in treating psoriasis vulgaris.

Effects of chinese formula jueyin granules on psoriasis in an animal model.

Although Traditional Chinese medicine (TCM) is known to be effective for psoriasis patients, the responsible mechanisms still remain poorly understood. In this study, we aimed to evaluate the effect of one formula, named Jueyin granules (JYG) in the mouse model of the vaginal epithelium and tail epidermis. Additionally, we also determined the anti-inflammatory effects of JYG in an imiquimod- (IMQ-) induced psoriasis-like skin mouse model. Our results show that JYG can attenuate the IMQ-induced psoriasis-like inflammation, accompanied with increased epidermal hyperplasia. We also measured estrogenic stage mitosis of vaginal epithelial cells and the formation of granular cell layers in male mouse tails per 100 scales, as well as the tissue nitric oxide (NO) and malondialdehyde (MDA) levels using the ELISA method. The results suggest that JYG significantly inhibited mitosis in mouse vaginal epithelial cells, promoted the formation of the squamous epidermal granular layer in mice tails, and reduced the levels of NO and MDA in an imiquimod-induced psoriasis-like skin mouse model after 14 d (P < 0.05). These results demonstrate that JYG might be an effective clinical treatment for psoriasis and the effects may be related to inhibited keratinocytes proliferation, improved parakeratotic epidermal cells, and reduced expression of NO and MDA.

[Effect of Shengji Huayu Recipe on the expression of MMP-3 and TIMP-1 in skin ulcer tissue of diabetic rats].

OBJECTIVE: To study the effect of Shengji Huayu Recipe (SHR)on the expression of MMP-3 and TIMP-1 in the skin ulcer tissue of diabetic rats. METHODS: The skin ulcer model was established in diabetic mice. Different compatibility proportions of SHR [the ratio of Shengji Recipe (SJR) to Huayu Recipe (HYR) = 2:1, 1:1, and 1:2, respectively] were used to intervene. The expression of MMP-3 protein in the skin ulcer of diabetic rats was detected by Western blot method,and TIMP-1 protein was detected by immunohistochemical assay. RESULTS: At each time point, there was no statistical difference in the blood glucose level among groups (P > 0.05). But all of them increased significantly,when compared with those of the normal wound group (P < 0.01). As for the difference between after would area treatment and before would area treatment, better effect was obtained in the SHR No. 3 group and the normal ulcer group than in the diabetic ulcer model group (P < 0.05). Results of Western blot showed that the MMP-3 protein expression was higher in the SHR No. 2 group than in the SHR No.3 group (P < 0.05). Immunohistochemical results showed that TIMP-1 protein expression was lower in the SHR No. 2 group than in the SHR No. 3 group and the diabetic ulcer model group (P < 0.05). TIMP-1 protein expression was higherin the SHR No. 3 group than in the SHR No. 2 group (P < 0.01). CONCLUSION: Using SHR No.3 was conducive to the promotion of wound healing in early wound repair stage, and using SHR No. 2 might be conducive to inhibiting the formation of pathological scar.

[Research on different expressions of peripheral blood Th1/Th2 cells in psoriasis patients of blood heat syndrome and of blood stasis syndrome].

OBJECTIVE: To explore the correlation between the pathogenesis of psoriasis patients of blood heat syndrome (BHS) and blood stasis syndrome (BSS) and peripheral blood Th1/Th2 cells axis drift, and to observe different expressions of peripheral blood Th1/Th2 cells between healthy subjects and psoriasis patients of BHS and BSS. METHODS: There were 15 patients in the BHS group and 15 in the BSS group. There were 16 patients in the healthy control group. The expressions of CD4+ gamma-interferon (IFN-gamma) and interleukin-4 (IL-4) in the peripheral blood mononuclear cells (PBMCs) were measured by flow cytometry (FACS). The Th1 main cytokines such as IFN-gamma and Th2 cytokines such as IL-4 in the serum of psoriasis patients of different syndromes were detected by enzyme-linked immunosorbent assay (ELISA). The Psoriasis Area and Severity Index score (PASI) were conducted. RESULTS: FACS results showed that the expression level of CD4+ IFN-gamma+ in the PBMCs was significantly higher in the BHS group than in the BSS group and the healthy control group (P < 0.05). Besides, it was positively correlated with the PASI (P < 0.05). ELISA results showed that the peripheral serum level of IFN-gamma was significantly higher in the BHS group than in the BSS group and the healthy control group (P < 0.05). The plasma level of IFN-gamma was positively correlated with the PASI score in the BHS group (P < 0.05). The plasma level of IFN-gamma was negatively correlated with the PASI score in the BSS group (P < 0.05). The peripheral serum level of IL-4 was significantly lower in the BHS group than in the BSS group and the healthy control group (P < 0.05). CONCLUSIONS: Peripheral Th1 cells had dominant state in psoriasis patients of BHS. When psoriasis patients of BHS were transformed to BSS or to the normal level, the expression of peripheral blood Th1 cells decreased.

Characteristics and clinical managements of chronic skin ulcers based on traditional chinese medicine.

Chronic skin ulcer (CSU), including diabetic ulcers, venous ulcers, radiation ulcers, and pressure ulcers, remains a great challenge in the clinic. CSU seriously affects the quality of life of patients and requires long-term dedicated care, causing immense socioeconomic costs. CSU can cause the loss of the integrity of large portions of the skin, even leading to morbidity and mortality. Chinese doctors have used traditional Chinese medicine (TCM) for the treatment of CSU for many years and have accumulated much experience in clinical practice by combining systemic regulation and tropical treatment of CSU. Here, we discuss the classification and pathogenic process of CSU and strategies of TCM for the intervention of CSU, according to the theories of TCM. Particularly, we describe the potential intervenient strategies of the "qing-hua-bu" protocol with dynamic and combinational TCM therapies for different syndromes of CSU.

Astragaloside IV Downregulates ß-Catenin in Rat Keratinocytes to Counter LiCl-Induced Inhibition of Proliferation and Migration.

Re-epithelialization is a crucial step towards wound healing. The traditional Chinese medicine, Astragalus membranaceus (Fisch) Bge, has been used for hundreds of years for many kinds of ulcerated wounds. Recent research has identified the active compound in this drug as astragaloside IV (AS-IV), but the underlying molecular mechanisms of its therapeutic action on keratinocytes remain poorly understood. In this study, we used an in vitro model of ulcer-like wound processes, lithium chloride (LiCl)-induced cultured mouse keratinocytes, to investigate the effects of AS-IV treatment. The effects on cell proliferation were evaluated by the MTS/PMS colorimetric assay, effects on cell migration were determined by a wound-healing scratch experiment, effects on the cell cycle were analyzed by flow cytometry, and effects on protein expression were analyzed by immunoblotting and immunofluorescence. LiCl strongly inhibited cell proliferation and migration, up-regulated ß-catenin expression, and down-regulated proliferating cell nuclear antigen (PCNA) expression. AS-IV treatment attenuat the inhibition of proliferation and migration, significantly reducing the enhanced ß-catenin expression, and recovering PCNA and ß-tubulin expression. Thus, AS-IV mediates mouse keratinocyte proliferation and migration via regulation of the Wnt signaling pathway. Down-regulating ß-catenin to increase keratinocyte migration and proliferation is one mechanism by which AS-IV can promote ulcerated wound healing.

Tanshinone IIA Inhibits Growth of Keratinocytes through Cell Cycle Arrest and Apoptosis: Underlying Treatment Mechanism of Psoriasis.

The aim of the present investigation was to elucidate the cellular mechanisms whereby Tanshinone IIA (Tan IIA) leads to cell cycle arrest and apoptosis in vitro in keratinocytes, the target cells in psoriasis. Tan IIA inhibited proliferation of mouse keratinocytes in a dose- and time-dependent manner and induced apoptosis, resulting in S phase arrest accompanied by down-regulation of pCdk2 and cyclin A protein expression. Furthermore, Tan IIA-induced apoptosis and mitochondrial membrane potential changes were also further demonstrated by DNA fragmentation, single-cell gel electrophoresis assay (SCGE), and flow cytometry methods. Apoptosis was partially blocked by the caspase-3 inhibitor Ac-DEVD-CHO. Mitochondrial regulation of apoptosis further downstream was investigated, showing changes in the mitochondrial membrane potential, cytochrome c release into the cytoplasm, and enhanced activation of cleaved caspase-3 and Poly ADP-ribose polymerase (PARP). There was also no translocation of apoptosis-inducing factor (AIF) from mitochondria to the nucleus in apoptotic keratinocytes, indicating Tan IIA-induced apoptosis occurs mainly through the caspase pathway. Our findings provide the molecular mechanisms by which Tan IIA can be used to treat psoriasis and support the traditional use of Salvia miltiorrhiza Bungee (Labiatae) for psoriasis and related skin diseases.

Effects of external application of Chinese medicine on diabetic ulcers and the expressions of ß-catenin, c-myc and K6.

OBJECTIVE: To observe the clinical efficacy of Chinese medicine (CM) treatment of Hongyou Ointment and Shengji Powder on diabetic ulcers, and to observe the influence of CM treatment on the expressions of proteins associated with the Wnt signaling pathway, such as ß-catenin, c-myc and K6. METHODS: sixty-two patients fitting the registration standards were randomly divided into the CM group (31 patients) and the Western medicine (WM) group (31 patients) by a random number table. The patients in the CM group were treated with Hongyou Ointment and Shengji Powder externally. The patients in the WM group were treated with mupirocin ointment, growth factor (bFGF), and Vaseline gauze for external use and with basic therapies. Wound-healing time and four-week healing rate were recorded. The wounds were measured by digital photography and ImageJ software. Skin biopsies were obtained from 24 patients before CM treatment and 20 patients after CM treatment. Immunohistochemical tests and semi-quantitative imaging with NIH ImageJ 1.42 software were used to analyze the changes in protein expression of ß-catenin, c-myc, and K6. RESULTS: Fifty-three patients completed the trial; four patients in the CM treatment group and five patients in the WM group dropped out. Among them, four were dissatisfied with the treatment process, two could not continue because of their jobs, two failed to complete the course of follow-up appointments, and one was diagnosed with squamous cell carcinoma during treatment. The comparison of ulcer healing rates between the two groups showed insignificant differences (P=0.77). The ulcer healing rates were 33.33% (9/27) in the CM group and 26.92% (7/26) in the WM group. However, the effective rate was significantly higher in the CM group (81.48%, 22/27) than in the WM group (57.69%, 15/26, P=0.04). The mean wound healing time was shorter in the CM group (22.71 ±5.46 days) than in the WM group (26.56 ±7.56 days, P=0.04). CM treatment was well tolerated, and there was no withdrawal due to adverse reactions. Immunohistochemical analysis in the refractory wound indicated higher expressions of ß-catenin, c-myc and K6 compared with the normal skin. ß-catenin was abnormally expressed in the nuclei of the keratinocytes and fibroblasts at the wound margins, and the expressions of c-myc and K6 were highly expressed in the full hyperplastic epidermis, especially in the granular layer (P<0.05). The expressions of these proteins decreased after CM treatment. The expression levels of ß-catenin, c-myc, and K6 proteins before and after the treatment were 101.88 ± 10.76 vs. 140.42 ±8.45; 113.27 ± 16.75 vs. 153.79 ±8.32; 90.39 ±11.07 vs. 151.29 ±7.39, respectively. CONCLUSIONS: CM treatment using Hongyou Ointment and Shengji Powder was efficient in the management of diabetic skin ulcers. The mechanism of action might be related to the inhibition of the Wnt signaling pathway.