Habitat selection by Asiatic black bear (Ursus thibetanus) in Siran and Kaghan Valleys, Pakistan / Seleção de habitat pelo urso-negro asiático (Ursus thibetanus) nos vales Siran e Kaghan, Paquistão

Asiatic black bear is present in variety of habitats like broad-leaves and coniferous forests, extending form sea level to 4300m elevation and change their habitat for food purpose seasonally. The present study was conducted at Kaghan and Siran Valleys, District Mansehra, Khyber Pakhtunkhwa, Pakistan to assess habitat of black bear. Line transect method was used for observation of bear signs. Twelve meter circular radius plots were selected for the concern vegetation’s i.e. (trees, shrubs and herbs) and three to six plots were placed in each transect. At the result of sign survey, thirteen different categories of bear signs were recorded and encounter rate was calculated for each sign. A total of 1858 signs were observed during field surveys. Total (81%) coniferous species were recoded among trees, with the highest appearance of Pinus wallichiana (34.22%) and Spruce spp (27.76%), similarly broad leaves trees (18.56%) were also recoded from habitat plots. Most of the signs were encountered in bushy areas, whereas high number of Viburnum Spp (60.29%) was present. It is indicated that black bear prefers blend of Coniferous Trees, Viburnum and Ferns Species; probably because these plants provide enough food, protection, and meticulous shelter because more than 80% of habitat composed of these three species. Currently habitat destruction and increase in human population are the up-growing issues for wild animals (especially Asiatic black bear), which is highly sensitive to such problems. High levels of conservation efforts are recommended for the protection of black bear habitat and to avoid human interference in their territory.

O urso-negro asiático está presente em uma variedade de habitats, como folhas largas e florestas de coníferas, estendendo-se do nível do mar até 4.300 m de altitude e mudando seu habitat para fins alimentares sazonalmente. O presente estudo foi conduzido nos vales Kaghan e Siran, distrito de Mansehra, Khyber Pakhtunkhwa, Paquistão, para avaliar o habitat do urso preto. O método de transecto linear foi usado para a observação de sinais do urso. Parcelas de raio circular de 12 m foram selecionadas para a vegetação de preocupação, ou seja, árvores, arbustos e ervas, e 3 a 6 parcelas foram colocadas em cada transecto. No resultado da pesquisa de sinais, 13 categorias diferentes de sinais de urso foram registradas e a taxa de encontro foi calculada para cada sinal. Um total de 1858 sinais foi observado durante os levantamentos de campo. O total (81%) de espécies de coníferas foi recodificado entre as árvores, com maior aparecimento de Pinus wallichiana (34,22%) e Spruce spp (27,76%); árvores de folhas largas (18,56%) também foram recodificadas em parcelas de habitat. A maioria dos sinais foi encontrada em áreas com arbustos, enquanto um alto número de Viburnum spp (60,29%) estava presente. É indicado que o urso-negro prefere a mistura de árvores coníferas, espécies de viburnos e samambaias; provavelmente porque essas plantas fornecem alimento suficiente, proteção e abrigo meticuloso porque mais de 80% do habitat é composto por essas três espécies. Atualmente, a destruição do habitat e o aumento da população humana são questões crescentes para os animais selvagens, especialmente o urso-negro asiático, que é altamente sensível a esses problemas. Altos níveis de esforços de conservação são recomendados para a proteção do habitat do urso-negro e para evitar a interferência humana em seu território.

Bear bile powder attenuates senecionine-induced hepatic sinusoidal obstruction syndrome in mice / 中国天然药物

Hepatic sinusoidal obstruction syndrome (HSOS) via exposure to pyrrolizidine alkaloids (PAs) is with high mortality and there is no effective treatment in clinics. Bear bile powder (BBP) is a famous traditional animal drug for curing a variety of hepatobiliary diseases such as cholestasis, inflammation, and fibrosis. Here, we aim to evaluate the protective effect of BBP against HSOS induced by senecionine, a highly hepatotoxic PA compound. Our results showed that BBP treatment protected mice from senecionine-induced HSOS dose-dependently, which was evident by improved liver histology including reduced infiltration of inflammatory cells and collagen positive cells, alleviated intrahepatic hemorrhage and hepatic sinusoidal endothelial cells, as well as decreased conventional serum liver function indicators. In addition, BBP treatment lowered matrix metalloproteinase 9 and pyrrole-protein adducts, two well-known markers positively associated with the severity of PA-induced HSOS. Further investigation showed that BBP treatment prevents the development of liver fibrosis by decreasing transforming growth factor beta and downstream fibrotic molecules. BBP treatment also alleviated senecionine-induced liver inflammation and lowered the pro-inflammatory cytokines, in which tauroursodeoxycholic acid played an important role. What's more, BBP treatment also decreased the accumulation of hydrophobic bile acids, such as cholic acid, taurocholic acid, glycocholic acid, as well. We concluded that BBP attenuates senecionine-induced HSOS in mice by repairing the bile acids homeostasis, preventing liver fibrosis, and alleviating liver inflammation. Our present study helps to pave the way to therapeutic approaches of the treatment of PA-induced liver injury in clinics.

Macroanatomic and morphometric analysis of the brown bear (Ursus arctos horribilis) mandible / Análisis macro anatómico y morfométrico de la mandíbula del oso pardo (Ursus arctos horribilis)

SUMMARY: Brown bear (Ursus arctos horribilis) is a wild animal from the bear (Ursidae) family. In this study, it was aimed to determine the morphometric values and anatomical structure of the brown bear mandible. After the superficial muscles of the mandible were dissected, the muscles were completely separated from the bones by boiling. 17 morphometric measurements were taken from the right and left mandible of each animal with the help of digital calipers. The mean and standard deviation values of the taken morphometric measurements were analyzed in the SPSS (20.0 version) package program. The mandible length was measured as 250.37 ± 9.75 mm on the right side and 246.83 ± 5.92 mm on the left side. The mandible height was determined as 105.76 ± 4.18 mm on the right and 108.62 ± 3.33 mm on the left. Consequently, the mandible was submitted to the results of the brown bear in the diversity of wildlife found in Turkey. We believe that the presented results will contribute to anatomical, surgical and archaeological studies.

RESUMEN: El oso pardo (Ursus arctos horribilis) es un animal salvaje de la familia de los osos (Ursidae). El objetivo de este estudio fue determinar los valores morfométricos y la estructura anatómica de la mandíbula del oso pardo, luego de la disección de los músculos superficiales de la mandíbula. Los músculos fueron separados por completo de los huesos mediante ebullición. Se tomaron 17 medidas morfométricas de la mandíbula derecha e izquierda de cada animal con la ayuda de calibradores digitales. Los valores de desviación estándar y media de las medidas morfométricas tomadas se anali- zaron en el programa SPSS (versión 20.0). El largo determiando de la mandíbula fue de 250,37 ± 9,75 mm en el lado derecho y 246,83 ± 5,92 mm en el lado izquierdo; la altura de la mandíbula era de 105,76 ± 4,18 mm en el lado derecho y 108,62 ± 3,33 mm en el izquierdo. Los resultados morfométricos obtenidos de la mandíbula del oso pardo en la diversidad de vida silvestre que se encuentra en Turquía contribuirán con el conocimiento anatómico y para los estudios quirúrgicos y arqueológicos.

Bear Bile Powder Inhibits Growth of Hepatocellular Carcinoma via Suppressing STAT3 Signaling Pathway in Mice / 中国结合医学杂志

OBJECTIVE@#To evaluate the inhibitory effect of bear bile powder (BBP) on hepatocellular carcinoma (HCC) growth in vivo and investigate the underlying mechanisms.@*METHODS@#A HCC xenograft mouse model was developed by producing with huh7 cells. After 5 days following xenograft implantation, ten HCC xenograft mice were given intra-gastric administration with 10 mg/(kg•d) dose of BBP or saline for 3 weeks. Tumor growth in HCC xenograft mice was evaluated by measuring the tumor weight and volume. Cell apoptosis, proliferation or tumor angiogenesis were examined via immunohistochemical (IHC) staining for transferase-mediated deoxyuridine triphosphate-biotin nick end labeling (TUNEL), proliferating cell nuclear antigen (PCNA) or cluster of differentiation 31 (CD31), respectively. Phosphorylation of signal transducer and activator of transcription 3 (STAT3) were determined by Western blot. The mRNA and protein expressions of Bcl-2, Bax, Cyclin D1 and Cyclin-dependent kinase 4 (CDK4) in HCC tumor tissues were respectively determined by reverse transcription polymerase chain reaction (RT-PCR) and Western blot. The protein expression of vascular endothelial growth factor A (VEGF-A) in tumor tissues was examined by IHC staining.@*RESULTS@#BBP treatment led to a significant decrease on tumor volume and tumor weight in HCC mice (P<0.05) and had no effect on the change of body weight. In addition, BBP profoundly promoted cell apoptosis, inhibited cell proliferation and intratumoral microvessel density in HCC tumor tissues (P<0.05). Moreover, BBP treatment remarkably suppressed the STAT3 phosphorylation and modulated the expression of critical target genes including Bcl-2, Bax, Cyclin D1, CDK4 and VEGF-A in HCC mice.@*CONCLUSION@#BBP exerts its anti-cancer activities via suppressing STAT3 signaling pathway and affecting multiple intracellular targets.

Genome misclassification of Klebsiella variicola and Klebsiella quasipneumoniae isolated from plants, animals and humans / Clasificación errónea de aislados de Klebsiella variicola y Klebsiella quasipneumoniae obtenidos de plantas, animales y humanos

Abstract: Objective: Due to the fact that K. variicola, K. quasipneumoniae and K. pneumoniae are closely related bacterial species, misclassification can occur due to mistakes either in normal biochemical tests or during submission to public databases. The objective of this work was to identify K. variicola and K. quasipneumoniae genomes misclassified in GenBank database. Materials and methods: Both rpoB phylogenies and average nucleotide identity (ANI) were used to identify a significant number of misclassified Klebsiella spp. genomes. Results: Here we report an update of K. variicola and K. Quasipneumoniae genomes correctly classified and a list of isolated genomes obtained from humans, plants, animals and insects, described originally as K. pneumoniae or K. variicola, but known now to be misclassified. Conclusions: This work contributes to recognize the extensive presence of K. variicola and K. quasipneumoniae isolates in diverse sites and samples.

Resumen: Objetivo: Identificar genomas mal clasificados de K. variicola, y K. quasipneumoniae en la base de datos del GenBank. Material y métodos: En el presente estudio se usaron tanto análisis filogenéticos usando rpoB como la identidad media de nucleótidos (ANI, por sus siglas en ingles) para identificar un número significativo de genomas del género Klebsiella. Resultados: Se reportó una actualización de genomas de K. variicola y K. quasipneumoniae correctamente clasificados y una lista de aquellos aislamientos obtenidos de seres humanos, plantas, animales e insectos, descritos originalmente como K. pneumoniae o K. variicola pero ahora se conoce que están mal clasificados. Conclusiones: Este trabajo contribuye a la presencia extensiva de aislamientos de K. variicola y K. quasipneumoniae en diversos sitios y muestras.

Identification of bear bile powder and its adulterants by using DNA barcoding technology / 中国中药杂志

To identify the precious bile powder and its adulterants by DNA barcoding, and establish its standard experimental process to ensure the safe and effective utilization. Total twelve sequences from samples of bear bile powder which come from Ursus thibetanus for DNA extraction, PCR(polymerase chain reaction) and sequence, then using CodonCode Aligner V 7.0.1 shear primer region to obtain COI sequence. The COI sequences of U. arctos and their adulterants were obtained from GenBank. MEGA7.0 software was applied for analyzing mutation, calculating intraspecific and interspecific K2P(Kimura 2-Parameter) genetic distance and constructing the Neighbor-joining tree(NJ). The results showed that the maximum K2P genetic distance of bear bile powder of U. thibetanus and U. arctos are far less than minimum K2P genetic distance within its adulterants species, and the results of NJ tree demonstrated that each species could be distinguished from the counterfeits obviously. DNA barcoding is a safe, convenient and reliable technique for species identification, and it is important to establish the standard sequence of COI sequences for animal medicines.

Study of characteristic ions in frequently-used traditional Chinese medicinal materials derived from animal bile and its application / 中国中药杂志

A LC-MS method was established for study of characteristic ions in frequently-used traditional Chinese medicinal materials derived from animal bile. UPLC-Q-TOF was used in the data acquisition work, then, the software of MarkerLynxTM v.4.1 was performed in the chemometric analysis of data. Besides, selected ion chromatograms of these bile acid ions were comparative studied. Better results were gained in the specificity identification of pig bile, bear bile, cultivated cow-bezoar , artificial cow-bezoar and some compound preparations(Hugan tablet and Rengongniuhuang Jiaxiaozuo capsule) by this method. The method is suitable for the specificity identification of pig bile, bear bile, cultivated cow-bezoar , artificial cow-bezoar and compound preparations containing these medicinal materials.

Intestinal rupture due to small bowel volvulus in a captive American black bear (Ursus americanus)

Intestinal rupture caused by small bowel volvulus was diagnosed in a captive 13-year-old male American black bear. The animal presented with decreased appetite, depression, lethargy, and mild abdominal distention for 3 days. The animal was treated with antibiotics, hypermetabolites, and digestive medicine daily; however, it died on the third day of treatment. The clinical symptoms included hemorrhagic ascites, gaseous extension of the small intestine, and intestinal rupture caused by small bowel volvulus. Hemorrhagic signs were observed in the lungs and heart. This is the first case to describe small bowel volvulus in mammals of the family Ursidae.

Bear bile powder inhibits angiogenesis in vivo and in vitro / 中国结合医学杂志

<p><b>OBJECTIVE</b>To evaluate the effect of bear bile powder (BBP) on angiogenesis, and investigate the underlying molecular mechanisms.</p><p><b>METHODS</b>A chick embryo chorioallantoic membrane (CAM) assay was used to evaluate the angiogensis in vivo. Human umbilical vein endothelial cells (HUVECs) were treated with 0, 0.25, 0.5, 0.75, and 1.0 mg/mL of BBP for 24, 48 and 72 h, respectively. The 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay was performed to determine the viability of HUVECs. Cell cycle progression of HUVECs was examined by fluorescence-activated cell sorting (FACS) analysis with propidium iodide staining. HUVEC migration was determined by wound healing method. An ECMatrix gel system was used to evaluate the tube formation of HUVECs. The mRNA and protein expression of vascular endothelial growth factor (VEGF)-A in both HUVECs and HepG2 human cells were examined by reverse transcription-polymerase chain reaction and enzyme linked immunosorbent assay, respectively.</p><p><b>RESULTS</b>Compared with the untreated group, BBP inhibited angiogenesis in vivo in the CAM model (P< 0.01). In addition, treatment with 0.25-1 mg/mL of BBP for 24, 48, or 72 h respectively reduced cell viability by 14%-27%, 29%-69% and 33%-91%, compared with the untreated control cells (P< 0.01). Additionally, BBP inhibited the proliferation of HUVECs via blocking the cell cycle G to S progression, compared with the S phase of untreated cells 48.05%± 5.00%, 0.25-0.75 mg/mL BBP reduced S phase to 40.38%± 5.30%, 36.54± 4.50% and 32.13± 3.50%, respectively (Pglt; 0.05). Moreover, BBP inhibited the migration and tube formation of HUVECs, compared with the tube length of untreated cells 100%± 12%, 0.25-0.75 mg/mL BBP reduced the tube length to 62%± 9%, 43%± 5% and 17%± 3%, respectively (p< 0.01). Furthermore, BBP treatment down-regulated the mRNA and protein expression levels of VEGF-A in both HepG2 cells and HUVECs.</p><p><b>CONCLUSION</b>BBP could inhibit the angiogenesis by reducing VEGF-A expression, which may, in part, explain its anti-tumor activity.</p>

Anti-atherosclerotic Effects of Bear Bile Powder in Shexiang Tongxin Dripping Pill: a Mechanism Study / 中国中西医结合杂志

<p><b>UNLABELLED</b>OBJECTIVE : To study the anti-atherosclerotic mechanism of bear bile powder (BBP) in Shexiang Tongxin Dripping Pill (STDP) , and to provide scientific evidence for treating atherosclerosis (AS) by its therapeutic characteristics of cool resuscitation.</p><p><b>METHODS</b>AS model was duplicated using ApoE-/- gene knocked mice fed with high-fat diet. Thirty ApoE-/- deficient male mice were divided into four groups according to body weight using random digit table, i.e., the model group (A, n =9), the STDP group (B, n=E7), the STDP without BBP group (C, n =7), and the BBP group (D, n =9). Besides, another 9 C57BL/6J male mice of the same age were recruited as a normal control group (E). All mice in Group B, C, and D were respectively administered with corresponding drugs (30, 30, and 0. 33 mg/kg) by gastrogavage. Equal volume of normal saline was administered to mice in Group A and E. All medication lasted for 8 successive weeks. Serum levels of inflammatory cytokines such as interleukin 2 (IL-2), interleukin 6 (IL-6), tumor necrosis factor a (TNF-α), interferon y (IFNγ), and oxidized low-density lipoprotein (ox-LDL) were measured by ELISA. Serum levels of malondialdehyde (MDA), activities of glutathione (GSH) and superoxide dismutase (SOD) were determined using biochemical assay. Contents of reactive oxygen species (ROS) in the aortic root was detected by dihydroethidum (DHE) fluorescent probe. Expression levels of microRNAs (such as miR-20, miR-21, miR-126, and miR-155) were detected by real-time PCR.</p><p><b>RESULTS</b>The fluorescence intensity of the aorta was obviously enhanced in Group A. But it was obviously attenuated in Group B, C, and D, and the attenuation was the most in Group B. Compared with Group E, serum levels of IL-2, IL-6, TNF-α, IFN-γ, oxLDL, and MDA all increased (P <0. 01), GSH contents and SOD activities decreased (P <0. 01), expression levels of miR-126, miR-21, and miR-155 in aorta increased (P <0. 01), and the expression level of miR-20 decreased in Group A (P<0. 01). Compared with Group A, serum levels of IL-2, IL-6, TNF-α, IFN-γ, oxLDL, and MDA were all down-regulated (P <0. 01), GSH contents and SOD activities were up-regulated (P <0. 01), expression levels of miR-126, miR-21, and miR-155 in aorta were down-regulated in Group B, C, and D (P <0. 01). The expression level of miR20 was up-regulated in Group B and D (P <0. 01). Compared with Group B, serum levels of IL-2, IL-6, TNF-α, IFN-γ increased (P <0.01); GSH contents and SOD activities decreased, levels of MDA and oxLDL increased (P <0. 01) in Group C and D. Expression levels of miR-20 and miR-155 were down-regulated in Group C and D (P <0. 01).</p><p><b>CONCLUSIONS</b>STDP played roles in significantly regulating inflammatory factors and oxidative stress factors. Its mechanism might be possibly associated with regulating expressions of miR-126, miR-21, miR-155, and miR-20 in aorta. BBP played significant roles in STDP.</p>

Expression of neuropeptide Y and long leptin receptor in gastrointestinal tract of giant panda / 生物工程学报

To study the expression and distribution of neuropeptide Y (NPY) and long leptin receptor (OB-Rb) in the gastrointestinal tract of giant panda, samples of three animals were collected from the key laboratory for reproduction and conservation genetics of endangered wildlife of Sichuan province, China conservation and research center for the giant panda. Paraffin sections of giant panda gastrointestinal tissue samples were observed using hematoxylin-eosin staining (HE) and strept actividin-biotin complex immunohistochemical staining (IHC). The results show that the intestinal histology of three pandas was normal and no pathological changes, and there were rich single-cell and multi-cell mucous glands, long intestinal villi and thick muscularis mucosa and muscle layer. Positive cells expressing NPY and OB-Rb were widely detected in the gastrointestinal tract by IHC methods. NPY positive nerve fibers and neuronal cell were widely distributed in submucosal plexus and myenteric plexus, especially in the former. They were arranged beaded or point-like shape. NPY positive cells were observed in the shape of ellipse and polygon and mainly located in the mucous layer and intestinal glands. OB-Rb positive cells were mainly distributed in the mucous layer and the laminae propria, especially the latter. These results confirmed that NPY and OB-Rb are widely distributed in the gut of the giant panda, which provide strong reference for the research between growth and development, digestion and absorption, and immune function.

Advances in studies on bear bile powder / 中国中药杂志

In this paper, a detailed analysis was made on relevant literatures about bear bile powder in terms of chemical component, pharmacological effect and clinical efficacy, indicating bear bile powder's significant pharmacological effects and clinical application in treating various diseases. Due to the complex composition, bear bile powder is relatively toxic. Therefore, efforts shall be made to study bear bile powder's pharmacological effects, clinical application, chemical composition and toxic side-effects, with the aim to provide a scientific basis for widespread reasonable clinical application of bear bile powder.

Echoes of silence: The unheard struggles of the physically impaired learners in the mainstream education

@#<p style="text-align: justify;">The inclusion of the physically impaired learners in the mainstream education has, lately, become a public concern as their manifold struggles will make or break their success academically. Despite this, there is a lack of action to address the problem as evidenced by the absence of a disability perspective in the Philippine context. This study, optimistically, endeavors to serve as one of the voices (amongst many) of the physically impaired. The objective is to further urge the authorities to take into consideration their pressing concerns.</p> <p style="text-align: justify;">This study utilized the descriptive phenomenological approach. The researcher employed purposive sampling with the following criteria: students from a public school who are in the mainstream; those who are willing to participate; are mentally sound and; those with consent from the schools they are attending and from their guardians. The researcher utilized interview to gather data from the respondents. Documentations and audio recording were used to facilitate the data gathering. This research has gained ethical approval from the UC-IERC.</p> <p style="text-align: justify;">Data were analyzed using the thematic analysis. Results revealed that the difficulties of the physically impaired learners vary and are dependent on the extent of their impairment. Exploring the verbatim accounts of the six (6) participants, the researcher extracted significant statements and organized the formalized meanings into clusters of themes. Three themes emerged from the data generated in the course of conversation with the physically impaired learners. The three themes are- "Cross to bear"; "Callous consideration", and; "Mind over body".</p> <p style="text-align: justify;">In conclusion, the participants were found to have common problems and difficulties with respect to their education. They struggled much with the physical structure of the school environment, inconsiderate classmates, lack of social acceptance, and the problem on the curriculum they have. As such, the researcher recommends that there shall be re-structuring of the guidelines or policies for the disabled students in the mainstream. With this, the physically impaired learners could have equal opportunities with the physically able. Further, the teachers and physically able students will be informed on how to help and accommodate children with disabilities. Because of the limitations of the study, the researcher recognizes that the findings may not be applied to other locale and to others with physical impairment. Thus, it is strongly recommended that a further study should be done considering a wider setting and population.</p>

Determination of tauroursodeoxycholic acid in compound bile capsule by HPLC / 中国中药杂志

<p><b>OBJECTIVE</b>To discriminate and determine of the artificial bear bile of the compound bile capsule.</p><p><b>METHOD</b>Taking the pharmacopoeia as reference, the artificial bear bile was discriminated and determined by HPLC.</p><p><b>RESULT</b>The compound bile capsule and the control sample had chromatographic peak at the same time from HPLC. The content of the artificial bear bile was above 10 mg per tablets.</p><p><b>CONCLUSION</b>The artificial bear bile of compound bile capsules can be discriminated effectively and determined accurately by HPLC method.</p>

Technological parameter optimization method for washing of coarse bear gall powder extracts based on NIR spectroscopy / 中国中药杂志

During the washing process of coarse bear gall powder extracts, it is necessary to adjust the amount of ethyl acetate according to the properties of raw materials, which aims to improving the yield and purity of the final product. In the research, using NIR spectra to reflect the comprehensive properties of coarse bear gall powder extracts, the process is optimized in a flexible way. Forty batches experiments are designed according to the weight ratio of ethyl acetate and coarse extracts of bear gall powder. The NIR spectra of the coarse extracts of bear gall powder are collected and processed using principal component analysis (PCA) method. The first 8 principal components combined with the amount of the ethyl acetate are used as the input variables, and calibration models are established to predict the yield and purity of the final product 30 batches are used as calibration set, which is used to establish the models, and other 10 batches are used as validation set, which is used for the performance appraisal of the established models. The correlation coefficients of the calibration, inner cross-validation and external validation for the purity model are 0.902, 0.896 and 0.883, respectively, and the RMSEC, RMSECV and RMSEP are 1.22%, 1.48% and 1.59%, respectively. The correlation coefficients of the calibration, inner cross-validation and external validation for the yield model are 0.921, 0.859 and 0.916, respectively, and the RMSEC, RMSECV and RMSEP are 1.39%, 1.65% and 1.53% respectively. This work demonstrated that NIR spectra combined with technology parameter could be used to predict the yield and purity of the final product. Using the established models, the most appropriate amount of the ethyl acetate can be determined according to the properties of the coarse bear gall powder extracts, and the yield and purity of the final product can be improved.

Bear bile powder (熊胆粉) induces apoptosis of human hepatocellular carcinoma cells via mitochondrion-dependent pathway / 中国结合医学杂志

<p><b>OBJECTIVE</b>To evaluate the effect of Bear Bile Powder(, BBP) on the growth and apoptosis of HepG2 human hepatocellular carcinoma cells, and investigate the possible molecular mechanisms mediating its anti-cancer activity.</p><p><b>METHODS</b>HepG2 cells were treated with 0.4-1.0 mg/mL of BBP for 24, 48 and 72 h. The viability of HePG2 cells was determined by MTT assay. Cellular morphology was observed via phase-contrast microscopy. Fluorescence-activated cell sorting analysis with Annexin-V/propidium idodide and 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethyl-benzimidazol-carbocyanine iodide (JC-1) staining was performed to determine cell apoptosis and the loss of mitochondrial membrane potential, respectively. Activation of caspase-9 and -3 was evaluated by a colorimetric assay.</p><p><b>RESULTS</b>The treatment with 0.4-1 mg/mL of BBP for 24, 48, or 72 h respectively reduced cell viability significantly by 7%-60%, 20%-90% or 25%-98%, compared with the untreated control cells (P<0.01). In addition, BBP treatment induced morphological changes in HepG2 cells. Furthermore, after treated with 0, 0.4, 0.6, 0.8 and 1.0 mg/mL of BBP, apoptosis cells (including early and late apoptotic cells) were 18.0%±1.3%, 34.9%±2.2%, 33.9%±2.8%, 37.4%±2.8% and 46.0%±2.5%, respectively (P<0.05); and the percentage of cells with reduced JC-1 red fluorescence were 6.6%±0.8%, 8.5%±0.8%, 13.5%±1.6%, 17.6%±2.3% and 46.7%±3.6%, respectively (P<0.01). Finally, BBP treatment significantly and dose-dependently induced activation of both caspase-9 and caspase-3 in HepG2 cells (P<0.05).</p><p><b>CONCLUSIONS</b>BBP could inhibit the growth of HepG2 hepatocellular cancer cells through mitochondrion-mediated apoptosis, which may, in part, explain its anti-cancer activity. BBP may be a potential novel therapeutic agent for the treatment of hepatocellular carcinoma.</p>

Effect of bear bile powder on STAT3 pathway in hepatocellular carcinoma xenograft / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To observe the effect of bear bile powder (BBP) on the STAT3 pathway and its downstream target genes of nude mice hepatocellular carcinoma (HCC) xenograft, and to explore its mechanism for treating HCC.</p><p><b>METHODS</b>The subcutaneous xenograft model was established using HepG2 cells. When the subcutaneous transplanted tumor was formed, naked mice were randomly divided into two groups, the BBP group and the control group. Mice in the BBP group were administered with BBP by gastrogavage, once daily for 3 consecutive weeks, while mice in the control group were administered with normal saline by gastrogavage, once daily for 3 consecutive weeks. The body weight and the tumor volume were measured once per week. By the end of medication, the tumor weight was weighed and the tumor inhibition ratio calculated. The apoptosis of the tumor tissue was detected by TdT-mediated dUTP nick end labeling (TUNEL). The expression of Bcl2-associated X protein (Bax), B cell lymphoma/eukemina-2 (Bcl-2), cyclin-dependent protein kinase (CDK4), cyclinD1 were detected by reverse transcription-polymerase chain reaction (RT-PCR). The protein expression levels of signal transducers and transcription activators 3 (p-STAT3), proliferating cell nuclear antigen (PCNA), Bax, Bcl-2, CDK4, and cyclinD1 were determined by immunohistochemistry.</p><p><b>RESULTS</b>BBP could inhibit the tumor volume and tumor weight, showing statistical difference when compared with the control group (P < 0.01). Results of TUNEL showed that BBP could significantly induce the apoptosis of hepatoma carcinoma cells. Results of RT-PCR showed that BBP could up-regulate the expression of Bax and down-regulate mRNA expression of Bcl-2, CDK4, and cyclinD1. Immunohistochemical results showed that BBP could up-regulate the expression of Bax and inhibit the protein expression of p-STAT3, PCNA, Bcl-2, CDK4, and cyclinD1.</p><p><b>CONCLUSION</b>BBP could induce the apoptosis of hepatoma carcinoma cells and inhibit their proliferation by regulating STAT3 pathway.</p>

Analysis on replacement of traditional Chinese medicine bear bile with bile acids based on drug properties / 中国中药杂志

<p><b>OBJECTIVE</b>To discuss the rationality of the clinical replacement of traditional Chinese medicine (TCM) bear bile with bile acid constituents, and analyze the difference between these constituents and bear bile in drug properties.</p><p><b>METHOD</b>Summarizing the drug properties of bear bile by reference to medical literatures for drug properties of TCM bear bile and Science of Traditional Chinese Medicine (China Press of Traditional Chinese Medicine, 2007). Analyzing and summarizing the pharmacological effects of main bile acid constituents according to relevant literatures for studies on pharmacological effects of main bile acid constituents in CNKI database. Predicating the drug properties of these bile acid constituents by using the drug property predication model established by the study group according the pharmacological effects of main bile acid constituents in the paper, and compare the prediction results with the drug properties of bear bile.</p><p><b>RESULT</b>Bile acid constituents in bear bile were mostly cold in property, bitter in taste, and the combination of their drug properties could reflect the combined drug properties of bear bile.</p><p><b>CONCLUSION</b>All of these bile acid constituents in bear bile could show part of effects of bear bile. Attention shall be given to regulate the medication scheme in clinical application according to actual conditions.</p>

Structure determination of three novel bile acids from bear bile powder / 药学学报

A method of LC-QTOF/MS combining with chemical synthesis has been used to determine the structures of three novel bile acids from bear bile powder. Reference substances of tauroursodeoxycholic acid and taurochenodeoxycholic acid were oxidized by pyridinium chlorochromate. The products were analyzed by LC-QTOF/MS. Total 4 products including 3 isomers were predicted and identified according to the PCC oxidation theory and LC-QTOF/MS results. Bear bile powder samples were dissolved by methanol and analyzed by LC-QTOF/MS. Three unknown peaks were found and identified as 2-[[(3beta, 5beta)-3-hydroxy-7, 24-dioxocholan-24-yl]amino]-ethanesulfonic acid, 2-[[(5beta)-3, 7, 24-trioxocholan-24-yl]amino]-ethanesulfonic acid and 2-[[(5beta, 7beta)-7-hydroxy-3, 24-dioxocholan-24-yl]amino]-ethanesulfonic acid, separately, by matching their results with that of oxidation products above.

Rapid identification of two new isomers in bear bile powder by LC-Q-TOF-MS combined with PCC oxidation / 中国中药杂志

A rapid method of Liquid chromatography-quadrupole time-of-flight mass spectrometry (LC-Q-TOF-MS) combined with pyridinium chlorochromate (PCC) oxidation has been developed to determine chemical structures of two novel isomers in bear bile powder. Derivatives of ursodeoxycholic acid (UDCA) and chenodeoxycholic acid (CDCA) were semi-synthesized by PCC oxidation, then were analyzed by LC-Q-TOF-MS. Separation was carried out on a reverse column with the mobile phase of acetonitrile-0.1% formic acid (45:55). The data of Q-TOF-MS was acquired by MS, MS/MS, positive and negative modes. Since UDCA and CDCA were stereochemical isomeric at an alcohol position, two oxidation products were same and have been confirmed by LC-Q-TOF-MS. Other two products were also determined based on the PCC oxidation theory. Samples of bear bile powder were dissolved by methanol and measured by LC-Q-TOF-MS. Two unknown peaks were found and identified by matching their retention times and accurate mass spectra ions with PCC oxidation productS. Finally, the structures of two new bile acids in bear bile powder were confirmed as 3alpha-hydroxy-7-oxo-5beta-cholanic acid, 7alpha-hydroxy-3-oxo-5beta-cholanic acid, respectively.