In vitro and in vivo antibacterial studies of volatile oil from Atractylodis Rhizoma against Staphylococcus pseudintermedius and multidrug resistant Staphylococcus pseudintermedius strains from canine pyoderma.

ETHNOPHARMACOLOGICAL RELEVANCE: Atractylodis Rhizoma is extensively employed in Traditional Chinese Medicine for the treatment of skin and gastrointestinal ailments. Its active components have been proven to demonstrate numerous beneficial properties, including antibacterial, antiviral, anti-inflammatory, anti-tumor, and anti-ulcer activities. Furthermore, the volatile oil from Atractylodis Rhizoma (VOAR) has been reported to effectively inhibit and eradicate pathogens such as Staphylococcus aureus, Escherichia coli and Candida albicans. Of particular concern is Staphylococcus pseudintermedius, the predominant pathogen responsible for canine pyoderma, whose increasing antimicrobial resistance poses a serious public health threat. VOAR merits further investigation regarding its antibacterial potential against Staphylococcus pseudintermedius. AIM OF THE STUDY: The study aims to verify the in vitro antibacterial activity of VOAR against Staphylococcus pseudintermedius. And a superficial skin infection model in mice was established to assess the in vivo therapeutic effect of VOAR. MATERIALS AND METHODS: Thirty strains of S. pseudintermedius were isolated from dogs with pyoderma, and the drug resistance was analyzed by disc diffusion method. The Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC) of VOAR were determined through the broth dilution method. The growth curve of bacteria in a culture medium containing VOAR was monitored using a UV spectrophotometer. Scanning electron microscopy was employed to observe the effects of VOAR on the microstructure of S. pseudintermedius. The impact of VOAR on the antibiotic resistance of S. pseudintermedius was assessed using the disc diffusion method. Twenty mice were randomly divided into four groups: the control group, the physiological saline group, the VOAR group, and the amikacin group. With the exception of the control group, the skin barrier of mice was disrupted by tap stripping, and the mice were subsequently inoculated with S. pseudintermedius to establish a superficial skin infection model. The modeled mice were treated with normal saline, VOAR, and amikacin for 5 days. Following the treatment period, the therapeutic effect of each group was evaluated based on the measures of body weight, skin symptoms, tissue bacterial load, tissue IL-6 content, and histopathological changes. RESULTS: The MIC and MBC of VOAR against 30 clinical isolates of S. pseudintermedius were found to be 0.005425% and 0.016875%, respectively. VOAR could exhibit the ability to delay the entry of bacteria into the logarithmic growth phase, disrupt the bacterial structure, and enhance the antibacterial zone in conjunction with antibiotic drugs. In the superficial skin infection model mice, VOAR significantly reduced the scores for skin redness (P < 0.0001), scab formation (P < 0.0001), and wrinkles (P < 0.0001). Moreover, VOAR markedly reduced the bacterial load (P < 0.001) and IL-6 content (P < 0.0001) in the skin tissues of mice. Histopathological observations revealed that the full-layer skin structure in the VOAR group was more complete, with clearer skin layers, and showed significant improvement in inflammatory cell infiltration and fibroblast proliferation compared to other groups. CONCLUSION: The results demonstrate that VOAR effectively inhibits and eradicates Staphylococcus pseudintermedius in vitro while also enhancing the pathogen's sensitivity to antibiotics. Moreover, VOAR exhibits a pronounced therapeutic effect in the superficial skin infection model mice.

Tetramethylpyrazine-Rhein Derivative inhibits the migration of canine inflammatory mammary carcinoma cells by mitochondrial damage-mediated apoptosis and cadherins downregulation.

BACKGROUND: Canine inflammatory mammary carcinoma (CIMC) has a high incidence of metastasis, high lethality, and poor prognosis, which needs novel adjuvant agents. Tetramethylpyrazine-Rhein Derivative (TRD) has been shown to have antitumor activity, which is a potential research direction for CIMC. PURPOSE: This study evaluated the efficacy of TRD on CIMC in vitro and in vivo, and provided possibilities for the application of active compounds in traditional Chinese medicine. METHODS: In vitro, TRD cytotoxicity was measured with CCK-8. Flow cytometry and transmission electron microscope were used to detect the cell cycle, cell death, and changes in mitochondria. Wound-healing assay, cell invasion assay, and scanning electron microscope were used to evaluate the suppression of cell migration and invasion. Expression changes were detected by RT-qPCR and western blot assay. In vivo, the lung metastasis models were randomly divided into control, low-dose TRD, high-dose TRD, and positive groups. Each group was administered orally once a day for 18 days and took in vivo imaging photos. RESULTS: The IC50 of TRD in CHMp and MDCK were 42.59 and 79.37 µM, respectively. TRD mediated cell apoptosis by mitochondrial damage and caused S and G2/M phase arrest by downregulating cyclin B1. Moreover, TRD reduced filopodia and inhibited cell migration by downregulating cadherins. In CIMC lung metastasis models, TRD could effectively inhibit tumor growth (P < 0.001) in the lungs without significant toxicity. CONCLUSION: TRD showed potential activity to inhibit CIMC lung metastasis with multi-target and low toxicity.

Evaluation of the antibacterial effect of Epigallocatechin gallate on the major pathogens of canine periodontal disease and therapeutic effects on periodontal disease mice.

Background: Periodontal disease (PD) is a prevalent oral affliction in canines, with limited therapeutic options available. The potential transmission of oral bacteria from canines to humans through inter-species contact poses a risk of zoonotic infection. Epigallocatechin gallate (EGCG), the principal catechin in green tea polyphenols, exhibits antibacterial properties effective against human PD. Given the clinical parallels between canine and human PD, this study explores the feasibility of employing EGCG as a therapeutic agent for canine PD. Methods and results: Initially, a survey and statistical analysis of bacterial infection data related to canine PD in China were conducted. Subsequently, the primary pathogenic bacteria of canine PD were isolated and cultivated, and the in vitro antibacterial efficacy of EGCG was assessed. Furthermore, verify the therapeutic effect of EGCG on a mouse PD model in vivo. The high-throughput 16S rRNA gene sequencing identified Porphyromonas, Fusobacterium, Treponema, Moraxella, and Capnocytophaga as the genera that distinguishing PD from healthy canines' gingival crevicular fluid (GCF) samples in China. The anaerobic culture and drug susceptibility testing isolated a total of 92 clinical strains, representing 22 species, from 72 canine GCF samples, including Porphyromonas gulae, Prevotella intermedia, Porphyromonas macacae, etc. The minimum inhibitory concentration (MIC) ranging of EGCG was from 0.019 to 1.25 mg/mL. Following a 7 days oral mucosal administration of medium-dose EGCG (0.625 mg/mL), the abundance of periodontal microorganisms in PD mice significantly decreased. This intervention ameliorated alveolar bone loss, reducing the average cementoenamel junction to the alveolar bone crest (CEJ-ABC) distance from 0.306 mm ± 0.050 mm to 0.161 mm ± 0.026 mm. Additionally, EGCG (0.3125 mg/mL) markedly down-regulated the expression of inflammatory factor IL-6 in the serum of PD mice. Conclusion: Our research demonstrates the significant antibacterial effects of EGCG against the prevalent bacterium P. gulae in canine PD. Moreover, EGCG exhibits anti-inflammatory properties and proves effective in addressing bone loss in a PD mouse model. These findings collectively suggest the therapeutic potential of EGCG in the treatment of canine PD. The outcomes of this study contribute valuable data, laying the foundation for further exploration and screening of alternative antibiotic drugs to advance the management of canine PD.

Tumor-associated macrophages in tumor progression and the role of traditional Chinese medicine in regulating TAMs to enhance antitumor effects.

Purpose: To emphasize the importance of tumor-associated macrophages (TAMs) in tumor immunity and to describe the ways in which extracts from Traditional Chinese Medicine (TCM) achieve tumor therapy by modulating macrophages. Significance: By summarizing these available data, this review focused on TAMs and TCM and can build the foundation for future research on antitumor therapeutics. Methods: In this review, we summarized the key functions of TAMs in cancer development and overviewed literature on TCM targeting TAMs together with other immune cells aiming to enhance antitumor immunity. Conclusions: With an indispensable role in antitumor immunity, TAMs contribute to tumor progression, migration, invasion, angiogenesis, lymphangiogenesis, and immunosuppressive microenvironment. In recent years, TCM has gradually gained attention as a potential antitumor adjunctive therapy in preclinical and clinical trials. TCM is also a regulator of cytokine secretion and cell surface molecule expression in balancing the tumor microenvironment (TME), especially macrophage activation and polarization. Therefore, it is believed that TCM could serve as modifiers with immunomodulatory capability.

Anti-inflammatory and anti-pruritic effects of Chi-Huang Solution in a murine model of allergic contact dermatitis.

ETHNOPHARMACOLOGICAL RELEVANCE: In treating atopic dermatitis, multi-mode management is adopted, including trying to avoid the allergens, controlling and preventing secondary infections, and using drugs to control itching. At present, most of the commonly used anti-pruritic drugs in the clinic are single-target and lead to serious side effects. Many studies have shown that a variety of traditional Chinese medicines have significant anti-inflammatory and anti-pruritic effects, and have the characteristics of multiple components, multiple targets, and multiple effects. AIM OF THE STUDY: The study aimed to explore the anti-inflammatory and anti-pruritic effects of the Chi-Huang Solution in a murine model of Allergic contact dermatitis (ACD). This study considers the effectiveness of the Chi-Huang Solution for external use on skin to provide an experimental basis for the clinical development and application of Chinese medicine and related preparations for Canine atopic dermatitis (CAD). MATERIALS AND METHODS: Forty-two male SPF C57BL/6 mice were randomly divided into control group (n = 6), ACD model group (n = 6), HAC control group (n = 6), and 4 Chi-Huang Solution groups (n = 6 in each group). With SADBE induce the murine model of ACD chronic pruritus, and initially evaluate whether the model is successful by counting scratching behavior, measuring the skin fold thickness and skin lesion score within 1 h. After treating the ACD model mice with deionized water, HAC, 1CH, 2CH, 3CH, and 4CH for 7 days, behavioral changes were used to evaluate the anti-pruritic effect. The skin fold thickness, skin lesion score, and spleen index were used to evaluate the anti-inflammatory effect of the Chi-Huang Solution. H.E. staining was used for the epidermal thickness measurement and pathological evaluation. RT-qPCR was used to analyze the mRNA expression of related inflammatory factors such as IL-1ß, TNF-α, IL-33, IL-4, IL-17A, CXCL10, and its receptor CXCR3 in the skin of the lesion site, as well as to detect the mRNA expression of pruritus-related genes such as TRPV1, TRPA1, and GRP in DRG. RESULTS: After the treatment of low-dose (0.1 g/mL) and medium-dose (0.2 g/mL) Chi-Huang Solution, the scratching times both decreased significantly (P < 0.05), meanwhile the medium-dose Chi-Huang Solution had an obvious effect on reducing scratches/scab score (P < 0.05). Moreover, no matter what dose it takes, all Chi-Huang Solution can alleviate the epidermal thickening (P < 0.05) and the infiltration of mast cells in the ACD murine model of ACD. It is worth mentioning that the count of mast cells in the dermis was significantly down-regulated after the treatment of medium-dose Chi-Huang Solution (P < 0.005). Furthermore, Chi-Huang Solution can significantly down-regulate the mRNA expression of related inflammatory factors in the skin, and reduce the mRNA expression of pruritus-related genes, such as TRPA1, TRPV1, and GRP in the spinal cord. CONCLUSIONS: The results indicated that Chi-Huang Solution for external use exhibits significant anti-inflammatory and anti-pruritic effects on SADBE-induced ACD chronic pruritus murine models. Chi-Huang Solution might emerge as an effective drug for the treatment of CAD and high-dose Chi-Huang Solution (0.4 g/ml) has better comprehensive effects.

Research Progress on the Mechanism of the Acupuncture Regulating Neuro-Endocrine-Immune Network System.

As one of the conventional treatment methods, acupuncture is an indispensable component of Traditional Chinese Medicine. Currently, acupuncture has been partly accepted throughout the world, but the mechanism of acupuncture is still unclear. Since the theory of the neuro-endocrine-immune network was put forward, new insights have been brought into the understanding of the mechanism of acupuncture. Studies have proven that acupuncture is a mechanical stimulus that can activate local cell functions and neuroreceptors. It also regulates the release of related biomolecules (peptide hormones, lipid hormones, neuromodulators and neurotransmitters, and other small and large biomolecules) in the microenvironment, where they can affect each other and further activate the neuroendocrine-immune network to achieve holistic regulation. Recently, growing efforts have been made in the research on the mechanism of acupuncture. Some researchers have transitioned from studying the mechanism of acupuncture as a single linear pathway to using systems approaches, including metabolomics, genomics, proteomics and biological pathway analysis. This review summarizes the research progress on the neuro-endocrine-immune network related mechanism of acupuncture and discusses its current challenges and future directions.

Integrative analyses identified ion channel genes GJB2 and SCNN1B as prognostic biomarkers and therapeutic targets for lung adenocarcinoma.

OBJECTIVES: Abnormal expressions of ion channel genes are associated with the occurrence and progression of tumors. At present, their roles in the carcinogenesis of lung adenocarcinoma (LUAD) are not clear. MATERIALS AND METHODS: Differentially expressed (DE) genes in the tumorigenesis were identified from 328 ion channel genes in 102 LUAD and paired adjacent normal samples. Similar analyses were performed between 177 metastatic and 286 non-metastatic LUAD samples to identify DE ion channel genes in the progression of LUAD. Independent prognostic factors selected from DE ion channel genes were used to construct a prognostic model. Correlation analysis and drugs-drug targets interaction network were used to screen the potential drugs for LUAD patients stratified by GJB2 or SCNN1B. RESULTS: Six ion channel genes (GJB2, CACNA1D, KCNQ1, SCNN1B, SCNN1G and TRPV6) were continuous differentially expressed in the tumorigenesis and progression of LUAD. The survival analysis in four datasets with 522 LUAD samples showed that GJB2 and SCNN1B were independent prognostic biomarkers. Patients with overexpression of GJB2 or underexpression of SCNN1B had shorter overall survival. Moreover, multi-omics analysis showed that hypomethylation of GJB2 and hypermethylation of SCNN1B in the promoter region may contribute to their aberrant expressions. KEGG enrichment analysis showed that the overexpressed genes in the group with high GJB2 or low SCNN1B were enriched in cancer-related pathways, while the underexpressed genes were enriched in metabolism-related pathways. The prognostic model with GJB2 and SCNN1B can stratify all LUAD patients into two groups with significantly different survival. Correlation analysis and drugs-drug targets interaction network suggested that GJB2 and SCNN1B expression might have indicative therapeutic values for LUAD patients. Finally, pan-cancer analysis in other eight cancer types showed that GJB2 and SCNN1B might be also potential prognostic factors for KIRC. CONCLUSIONS: GJB2 and SCNN1B were identified as prognostic biomarkers and therapeutic targets for LUAD.

Apigenin enhances viability of random skin flaps by activating autophagy.

Random skin flap is widely used in plastic surgery. However, flap necrosis caused by ischemia-reperfusion injury limits its clinical applications. Apigenin, a naturally occurring flavonoid mainly derived from plants, facilitates flap survival. In this study, we explored the effects of apigenin on flap survival and the underlying mechanisms. A total of 54 mice having a dorsal random flap model were randomly divided into control, apigenin, and apigenin +3-methyladenine groups. These groups were treated with dimethyl sulfoxide solution, apigenin, and apigenin +3-methyladenine, respectively. The animals were then euthanized to assess angiogenesis, apoptosis, oxidative stress, and autophagy levels through histological and protein analyses. Apigenin promotes survival of the skin flap area and reduces tissue edema. In addition, apigenin enhanced angiogenesis, attenuated apoptosis, alleviated oxidative stress, and activated autophagy. Interestingly, 3-methyladenine reversed the effects of apigenin on flap survival, angiogenesis, apoptosis, and oxidative stress through inhibition of autophagy. The findings of this study show that apigenin promotes angiogenesis, inhibits cell apoptosis, and lowers oxidative stress by mediating autophagy, thus the improving survival rate of random skin flaps.

Acupuncture Combined with Chinese Medicine Iontophoresis Treatment for Chronic Progressive Cervical Intervertebral Disk Disease in a Dog.

CASE REPORT: A 12-year-old castrated male dog with nonambulatory tetraplegia was diagnosed with spinal stenosis at C3-C4 through X-ray examination and with ventral extradural spinal compression at C3-C4 through myelography and computed tomography. The diagnosis of traditional Chinese veterinary medicine was local Qi and blood stagnation, spleen Qideficiency, blood deficiency, and kidney Yang deficiency. We initiated treatment using a combination of acupuncture and Chinese medicine iontophoresis with laser therapy. After 12 treatment days, there was a significant improvement in the dog's ambulation function, which was indicated by proper walking and flexible head-turning. CONCLUSION: This indicates that combining acupuncture and Chinese medicine iontophoresis could be a potential treatment for chronic progressive cervical intervertebral disk disease in dogs.

Simultaneous determination of gastrodin and puerarin in rat plasma by HPLC and the application to their interaction on pharmacokinetics.

Gastrodin (Gas) and puerarin (Pur) are bioactive substances derived from traditional Chinese medicine Gastrodia elata and Radix Puerariae, respectively, which were often used together in Chinese clinical prescriptions. Their injections were used in combined way for treatment of some cardiocerebrovascular diseases in clinic, especially for vertigo due to vertebrobasilar ischemia. In this paper, interaction of gastrodin and puerarin in rat plasma pharmacokinetics via intragastic (i.g.)/intravenous (i.v.) administration was investigated. A reliable HPLC method was developed for simultaneous determination of Gas and Pur in rat plasma with a linear range of 0.101-101 µg/mL for Gas and 0.0500-5.98 µg/mL for Pur (r(2)>0.993). The LLOQ, LOD of Gas and Pur were determined to be 0.101, 0.0486 µg/mL, and 0.05, 0.0245 µg/mL, respectively. The intra-day and inter-day precision were all less than 12.0%, whilst the accuracy were all within 96.4±6.00%. The proposed method has been successfully applied to the pharmacokinetic study of the analytes in rats after i.g./i.v. administration of Gas and Pur alone or combined with each other (i.g.: 40 mg/kg Gas, 400 mg/kg Pur; i.v.: 20 mg/kg Gas, 20 mg/kg Pur). Blood samples were collected from retinal vein plexus of rats at predetermined time points and plasma containing the internal standard tyrosol (IS) were precipitated by methanol and chromatography was carried out on a C(18) column with a gradient mobile phase of ACN-H(2)O with 0.05% phosphoric acid as a modifier. The pharmacokinetic profiles of combined administration were found to be distinct from those of given alone. The C(max), T(max), T(1/2), MRT of Gas administrated alone or combined with Pur via i.g. were 21.7 µg/mL, 0.250 h, 2.81 h, 0.830 h and 18.4 µg/mL, 0.550 h, 0.970 h, 1.37 h, respectively, of Pur administrated alone or combined with Gas via i.g. were 0.490 µg/mL, 1.95 h, 1.33 h, 2.10 h and 2.01 µg/mL, 0.570 h, 4.00 h, 5.10 h, respectively. The relative oral bioavailability of Pur in combined administration was 10.7 times as much as that of single administration, whilst 1.52 folds in Gas. These results indicate that co-administration of Gas and Pur is a promising combination to gain higher bioavailability and it is suggested that doctors pay more attention to the dosages of the two when simultaneously using both of them.