Quantification of Chemical Groups and Quantitative HPLC Fingerprint of Poria cocos (Schw.) Wolf.

(1)Objective: In this study, a quantitative analysis of chemical groups (the triterpenoids, water-soluble polysaccharides, and acidic polysaccharides) and quantitative high liquid performance chromatography (HPLC) fingerprint of Poria cocos (Schw.) Wolf (PC) for quality control was developed. (2) Methodology: First, three main chemical groups, including triterpenoids, water-soluble polysaccharides, and acidic polysaccharides, in 16 batches of PC were evaluated by ultraviolet spectrophotometry. Afterward, the quantitative fingerprint of PC was established, and the alcohol extract of PC was further evaluated. The method involves establishing 16 batches of PC fingerprints by HPLC, evaluating the similarity of different batches of PC, and identifying eight bioactive components, including poricoic acid B (PAB), dehydrotumulosic acid (DTA), poricoic acid A (PAA), polyporenic acid C (PAC), 3-epidehydrotumulosic acid (EA), dehydropachymic acid (DPA), dehydrotrametenolic acid (DTA-1), and dehydroeburicoic acid (DEA), in PC by comparison with the reference substance. Combined with the quantitative analysis of multi-components by a single marker (QAMS), six bioactive ingredients, including PAB, DTA, PAC, EA, DPA, and DEA, in PC from different places were established. In addition, the multivariate statistical analyses, such as principal component analysis and heatmap hierarchical clustering analysis are more intuitive, and the visual analysis strategy was used to evaluate the content of bioactive components in 16 batches of PC. Finally, the analysis strategy of three main chemical groups in PC was combined with the quantitative fingerprint strategy, which reduced the error caused by the single method. (3) Results: The establishment of a method for the quantification of chemical groups and quantitative HPLC fingerprint of PC was achieved as demonstrated through the quantification of six triterpenes in PC by a single marker. (4) Conclusions: Through qualitative and quantitative chemical characterization, a multi-directional, simple and efficient routine evaluation method of PC quality was established. The results reveal that this strategy can provide an analytical method for the quality evaluation of PC and other Chinese medicinal materials.

Saikosaponin-d Inhibits the Hepatoma Cells and Enhances Chemosensitivity Through SENP5-Dependent Inhibition of Gli1 SUMOylation Under Hypoxia.

Chemosensitivity is one of the key factors affecting the therapeutic effect on cancer, but the clinical application of corresponding drugs is rare. Hypoxia, a common feature of many solid tumors, including hepatocellular carcinoma (HCC), has been associated with resistance to chemotherapy in part through the activation of the Sonic Hedgehog (SHh) pathway. Hypoxia has also been associated with the increased SUMOylation of multiple proteins, including GLI family proteins, which are key mediators of SHh signaling, and has become a promising target to develop drug-resistant drugs for cancer treatment. However, there are few target drugs to abrogate chemotherapy resistance. Saikosaponin-d (Ssd), one of the main bioactive components of Radix bupleuri, has been reported to exert multiple biological effects, including anticancer activity. Here, we first found that Ssd inhibits the malignant phenotype of HCC cells while increasing their sensitivity to the herpes simplex virus thymidine kinase/ganciclovir (HSVtk/GCV) drug system under hypoxia in vitro and in vivo. Furthermore, we had explored that GLI family activation and extensive protein SUMOylation were characteristics of HCC cells, and hypoxia could activate the SHh pathway and promote epithelial-mesenchymal transition (EMT), invasion, and chemosensitivity in HCC cells. SUMOylation is required for hypoxia-dependent activation of GLI proteins. Finally, we found that Ssd could reverse the effects promoted by hypoxia, specifically active sentrin/small ubiquitin-like modifier (SUMO)-specific protease 5 (SENP5), a SUMO-specific protease, in a time- and dose-dependent manner while inhibiting the expression of SUMO1 and GLI proteins. Together, these findings confirm the important role of Ssd in the chemoresistance of liver cancer, provide some data support for further understanding the molecular mechanisms of Ssd inhibition of malignant transformation of HCC cells, and provide a new perspective for the application of traditional Chinese medicine in the chemical resistance of liver cancer.

Ligation of thoracic duct during thoracoscopic esophagectomy can lead to decrease of T lymphocyte.

BACKGROUND: Video-assisted thoracoscopic esophagectomy has been one of the most preferable surgical treatments for early esophageal cancer. Some scholars suggested that the thoracic duct should be routinely ligated to reduce the incidence of postoperative chylothorax, while another group raised an objection. As a classic indicator of immune function, T lymphocyte subsets can be applied to assess the effects of prophylactic thoracic duct ligation during thoracoscopic esophagectomy. METHODS: A total of 60 patients were recruited and randomized into thoracic duct ligation group and nonligation group. Venous blood was collected before and after video-assisted esophagectomy. The lymphocyte count and percentage, T lymphocyte subsets percentage were measured with fully automatic hemacytometer analyzer and flow cytometry. The difference between two groups was compared with t-test and the classified data were compared with Chi-square test. RESULTS: No significant difference was observed in peripheral blood CD3+, CD3+CD4+, and CD3+CD8+ lymphocyte percentage between the two groups before operation (P > 0.05). The mean value of peripheral blood CD3+, CD3+CD4+ lymphocyte percentage in ligation group was obviously less than that of in nonligation group after operation (P < 0.05). The mean of CD3+CD8+ lymphocyte percentage in ligation group was obviously higher than that of in nonligation group after operation (P < 0.05). CONCLUSION: Ligation of thoracic duct during esophagectomy could lead to decreased percentage of T lymphocyte and CD4+ Tlymphocyte, especially after arch of azygos vein had been transected. The thoracic duct should be selectively ligated during esophagectomy.

B-cell specific Moloney leukemia virus insert site 1 and peptidyl arginine deiminase IV positively regulate carcinogenesis and progression of esophageal squamous cell carcinoma.

High expression of B-cell specific Moloney leukemia virus insert site 1 (Bmi-1) and peptidyl arginine deiminase IV (PADI4) is associated with esophageal carcinoma. However, few studies have investigated the association between the Bmi-1 and PADI4 genes. The aim of the present study was to evaluate the expression of Bmi-1 and PADI4 and identify the association between the Bmi-1 and PADI4 genes in esophageal squamous cell carcinoma (ESCC) tissues. Bmi-1 and PADI4 gene expression levels were measured using immunohistochemistry, western blotting and reverse transcription-quantitative polymerase chain reaction in ESCC tissues from 86 patients who had not received pre-operative chemoradiation. The results revealed that the Bmi-1 and PADI4 genes had increased expression in carcinoma tissues compared with pericarcinous tissue (P<0.05). Bmi-1 gene expression was revealed to be associated with differentiation degree, clinical stage and lymph node metastasis (P<0.05), but had no association with gender, age or depth of invasion (P>0.05). The expression of PADI4 was associated with clinical stage, depth of invasion and lymph node metastasis (P<0.05), but was not associated with gender, age or differentiation degree (P>0.05). In addition, there was a positive association between Bmi-1 and PADI4 gene expression in ESCC (P<0.05). These results indicated that Bmi-1 and PADI4 positively regulate carcinogenesis and progression of ESCC.

Effect of Ligation of the Thoracic Duct During Oesophagectomy on the Absorption of D-xylose.

OBJECTIVE: To assess if prophylactic thoracic duct ligation during oesophagectomy influences the absorptive function of oesophageal cancer patients. STUDY DESIGN: Randomized controlled trial. PLACE AND DURATION OF STUDY: Department of Thoracic Surgery, Tai'an City Central Hospital, Tai'an, from August 2014 to December 2015. METHODOLOGY: Based on the management of the thoracic duct during oesophagectomy, 60 patients were randomized into two groups. D-xylose absorption test was used to evaluate the absorptive function. The two-independent-samples t-test was employed for statistical analysis with statistical significance at p < 0.05. RESULTS: The serum D-xylose concentration of ligation-group was significantly lower than that of no-ligation group on the first day after operation, (t=2.82, p=0.0066). However, there was no significant differences between them even before operation (t=1.34, p=0.1849). CONCLUSION: Ligation of the thoracic duct during oesophagectomy immediately affected the absorption of D-xylose, which may lead to malabsorption in the long run.

A giant infiltrating angiolipoma of the mediastinum: a case report.

BACKGROUND: Angiolipoma is a rare benign neoplasm composed of mature fatty tissue and multiple small abnormal blood vessels. Infiltrating mediastinal angiolipoma is an extremely rare tumor associated with delayed diagnosis. CASE PRESENTATION: A 42-year-old woman was presented with 3-month history of mild chest tightness. Imaging of the chest showed a large mass with fat densities in the middle superior mediastinum. A presumptive diagnosis was a tumor of liposarcoma. The patient was scheduled for a thoracotomy. After the excision, the symptoms were relieved and histological study revealed that the tumor was an angiolipoma. The patient recovered very well and was discharged 7 days after the surgery. After 7 months of follow-up the patient was clinically well and asymptomatic. CONCLUSIONS: We described a giant infiltrating mediastinal angiolipoma and its removal, and discussed the tumor characteristics and prognosis. Although extremely rare, infiltrating angiolipoma should be considered in the differential diagnosis of mediastinum lesions. The prognosis after surgical management of our patient is favorable.

Synchronous double primary cancer of the lung and nasal vestibule: A case report and literature review.

Squamous cell carcinoma (SCC) of the nasal vestibule is a rare tumor entity, and its occurrence combined with lung cancer is even rarer. Thus, several patients are often initially misdiagnosed or remain undiagnosed. This is the case report of a 55-year-old male patient who presented to our hospital with a neoplasm in the left lung. The patient was treated with left upper pulmonary lobectomy and the subsequent histopathological examination of the surgical specimen revealed a poorly differentiated SCC. On postoperative week 4, the patient presented with purulent and bloody discharge from the left nostril and was misdiagnosed with an upper jaw cyst. After another 3 weeks, the patient was re-admitted to the hospital with a mass of left nostril and nasal congestion. Tru-Cut biopsies from the nasal area and histopathological examination revealed a moderately differentiated SCC. According to the clinical presentation and the histopathological findings, the patient was diagnosed with double primary cancer of the lung and the nasal vestibule. The mass of the left nostril was significantly reduced in size with radiotherapy. To the best of our knowledge, there is no similar case previously reported in the literature. Due to the rarity of scc of the nasal vestibule concomittant with lung cancer, we herein present this case report with a review of the relevant literature and investigation of the clinical characteristics.

Treatment of esophageal-gastric double primary cancer by pedunculated remnant gastric interposition, esophageal-gastric anastomosis and gastrojejunal Billroth II anastomosis: A case report.

With the continuous advancement of clinical diagnostic techniques, including imaging technology, the incidence of confirmed multiple primary cancers or double primary carcinoma increases yearly. However, studies reporting synchronization surgery performed for primary dual esophageal gastric cancer are rare. The present study reports the case of a patient with double primary esophageal-gastric cancer, located in the thoracic cavity segment of the esophagus and gastric antrum of the stomach, respectively. The gastric cancer was diagnosed by endoscopy biopsy with concomitant esophageal cancer. The patient underwent gastric cancer resection, and pedunculated remnant gastric interposition esophagogastric side anastomosis was performed with gastrojejunostomy Billroth II anastomosis behind the colon. Abdominal cavity lymph node dissection was also performed. The esophageal-gastric double primary cancer was simultaneously excised and the gastric regions were used in the construction of the upper gastrointestinal tract: The surgery was successful. However, two weeks after surgery, upper gastrointestinal imaging revealed esophagogastric anastomotic leakage. Subsequently, an esophageal stent was inserted and antibiotics and additional treatment was administered. Follow-up one year after surgery revealed that the patient was well and remained in a stable condition.

Clinical Application of Recombinant Human Endostatin in Postoperative Early Complementary Therapy on Patients with Non-small Cell Lung Cancer in Chinese Mainland.

OBJECTIVE: To explore the clinical application of recombinant human endostatin (Endostar) in the treatment of patients with non-small cell lung cancer (NSCLC) in Chinese mainland. MATERIALS AND METHODS: A total of 75 patients diagnosed as NSCLC were randomly divided into control group (37 cases) and treatment group (38 cases). Control group was treated with postoperative complementary chemotherapy containing two-agent platinum protocol on postoperative d21, 3 weeks as a cycle, for totally 4~6 cycles. On this basis, treatment group was added with Endostar 7.5 mg/m2 on postoperative d8~9, 3~4 h/time, qd, 14 weeks as a cycle, for totally 4 cycles. The interval between every two cycles was 7 d. The 5-year progression-free survival (PFS), 5-year survival time and complications in both groups were observed. RESULTS: Compared with control group, the average PFS increased evidently in treatment group by 9.8 months (41.6 months vs. 31.8 months), and there was significant difference (P<0.05). And the median PFS was 42.5 months in treatment group, obviously longer than that in control group (33.7 months) by 8.8 months (P<0.05). Additionally, the 5-year overall survival rate (OS), average survival time and median survival time (MST) were 47.4%, 50.1 months and 59.3 months in treatment group, significantly higher than the 29.7%, 42.1 months and 43.5 months in control group (P<0.05). Only 1 patient showed poor healing of surgical wound in treatment group, but no surgery-associated complication was found in control group. Moreover, the postoperative complementary therapy-connected complication rates were 63.2% (24/38) and 59.5% (22/37) in treatment group and control group respectively, but there was no significant difference (P>0.05). CONCLUSIONS: The application of Endostar combined with sensitive platinum-contained chemotherapeutic agents in the postoperative complementary chemotherapy can be widely used in clinic because it can significantly prolong the long-term survival time of patients with NSCLC.

[Clinical application of a fully covered self-expandable metallic stent in treatment of airway fistula].

OBJECTIVE: To assess the feasibility, safety and efficacy of the use of a fully covered self-expandable stent for the treatment of airway fistula. METHODS: From August 2005 to November 2011, 9 patients underwent treatment by the introduction of a tracheo-bronchial or bronchial fully covered self-expandable metallic stent. There were 7 males and 2 females, aged from 28-65 years with a mean of 46 years. In this group, 7 cases were diagnosed as bronchopleural fistula, 1 case as tracheopleural fistula, 1 case as broncho-esophageal fistula, 8 cases with thoracic empyema. The fistula orifices were from 3.5-25.0 mm in diameter with a mean 8.4 mm. All patients received topical anesthesia, and L-shaped stent was placed in 6 patients and I-shaped stent in 3 patients under fluoroscopic guidance. After the stent placement, the patients with empyema were treated with continual irrigation of the empyema cavity. RESULTS: Stent placement in the tracheo-bronchial tree was technically successful in all patients, without procedure-related complications. The operating time was from 5-16 minutes, mean time (10 ± 4) minutes. Except for 1 patient, immediate closure of the airway fistula was achieved in the other patients after the procedure, as shown by the immediate cessation of bubbling in the chest drain system or the contrast examination. In this study, 1 patient coughed the inserted stent out due to irritable cough on the 5th day and had to receive repositioning of a new stent. Among the patients who were with empyema, 1 patient died of septicemia on the 8th day and 1 patient died of brain metastases from lung cancer 6 months after the stent insertion with empyema not cured, the other 6 patients' empyema healed from 2-5 months, mean time 3.7 months. Seven patients were followed from 3 to 36 months with a median of 13.5 months. During follow-up, 1 stent was removed from a patient 8 months after the stent implantation without empyema recurred. The remaining patient presented good tolerability to the existence of stent. The stents remained stable, no migration occurred, no empyema recurred, and the patient with broncho-esophageal fistula fed and drunk well. CONCLUSION: The use of fully covered self-expandable stents proved to be a safe, effective and fast minimally invasive method to treat airway fistula, especially for patients with a higher surgical risk or other failed treatments.

[Inhibitory effect of folic acid/polyamide-amine as a miR-7 vector on the growth of glioma in mice].

OBJECTIVE: To explore if folic acid/polyamide-amine (FA/PAMAM) enhances the therapeutic effect of miR-7gene therapy for glioma in vivo. METHODS: The miR-7 gene was transfected into U251 glioma cells by FA/PAMAM. The efficiency of gene transfection was assessed by fluorescence microscopy. The miR-7 level was detect by quantitative RT-PCR. Intracranial glioma models were established in thymectomized mice, and FA/PAMAM nanoparticles were transplanted into the tumors in situ 3 days later. The animal survival was recorded and the gross tumor volume and degree of edema were observed by MRI. Apoptosis in the glioma cells and expression of proliferating cell nuclear antigen (PCNA), matrix metalloproteinases 2 and 9 (MMP-2 and MMP-9) were assessed by immunohistochemistry, and EGFR and AKT-2 protein expression was detected by Western blot assay. RESULTS: Compared with the liposomes, the FA/PAMAM nanoparticles were more efficient to transfer miR-7 gene into U251 glioma cells, MRI showed that the tumor growth was much slower in the FA/PAMAM/miR-7 group, and the animal survival time was longer. The apoptosis rate was (5.3 ± 0.9)% in the control group, (11.4 ± 2.4)% in the liposome/miR-7 group, and (17.7 ± 3.7)% in the FA/PAMAM/miR-7 group. The immunohistochemical assay showed that the levels of PCNA, MMP-2 and MMP-9 protein in the FA/PAMAM/miR-7 group were (34.6 ± 5.4)%, (24.5 ± 4.1)%, (25.4 ± 5.1)%, respectively, significantly lower than those in the liposome/miR-7 group (49.3 ± 5.9)%, (31.7 ± 7.1)% and (39.4 ± 6.4)%, respectively, and those in the control group (57.3 ± 7.4)%, (45.4 ± 6.9)% and (55.1 ± 7.3)%, respectively (all P < 0.05). The expressions of EGFR and AKT-2 proteins were 1.09 ± 0.12 and 0.62 ± 0.10 in the control group, 0.63 ± 0.11 and 0.43 ± 0.07 in the liposome/miR-7 group, and significantly deceased (0.47 ± 0.09 and 0.31 ± 0.04, respectively) in the FA/PAMAM/miR-7 group (all P < 0.05). CONCLUSION: Compared with the liposomes, FA/PAMAM can transfect miR-7 into glioma cells with a higher efficiency in vivo, makes a longer time of the drug action, and shows a certain inhibitory effect on the growth of glioma, therefore, might become a new drug targeting agent in gene therapy forglioma.

[Study on reversion of malignant phenotype of glioma by siRNA targeting p75 neurotrophin receptor].

OBJECTIVE: To study the therapeutic efficacy of siRNA fragments silencing p75 neurotrophin receptor (p75(NTR)), which may be a key regulator of glioma cell apoptosis and invasion. METHODS: The siRNA sequence fragments targeting p75(NTR) were designed and transferred into human glioma cell line U251. RT-PCR and immunocytochemistry method were used to explore the expression of p75(NTR) mRNA and protein. Cell adhesion assay was employed to detect cellular adhesion ability, and soft agar clone formation assay was adopted to identify oncogenicity, and a U251 glioma model was established in nude mice. The intracranial tumor volume was detected by MRI. The expression of p75(NTR), NGF and cyclin D2 were identified using immunohistochemistry. Cell apoptosis was detected by apoptosis kit in situ. RESULTS: The siRNA fragments targeting p75(NTR) were capable of decreasing mRNA and protein expression of p75(NTR) in U251 glioma cell line. Both the cellular adhesion ability and oncogenicity were weakly relevant. The p75(NTR) expression level was negatively correlated with cyclin D2 and apoptosis, and positively correlated with NGF expression. The siRNA sequence fragments targeting p75(NTR) were effective in decreasing the gross volume of tumor; prolonged the survival time of mice, and the edge of tumor was much sharper than that of the control group. CONCLUSIONS: The gene silencing technique by siRNA targeting p75(NTR) is capable of decreasing tumor invasion and cell proliferation as well as inducing cell apoptosis. It is expected to be a new choice for glioma gene therapy.

[Differential proteomics in glioblastoma].

OBJECTIVE: To establish differential proteomics profiles of glioblastoma cell lines from Chinese, and to provide reference for future basic studies. METHODS: Total protein was extracted from 3 glioblastoma cell lines, CHG-5, TJ899 and TJ905. After normalization, the total protein was presented by two-dimensional (2D) electrophoresis, scanned and analyzed. Some of the identified protein spots were verified by immunocytochemistry of cell lines and immunohistochemistry of solid tumors. The glia cells were used as the control throughout the study. RESULTS: A total of 13 differential protein spots were selected, and eventually 10 were identified as unique proteins. These 10 proteins were involved in cytoskeleton forming, cellular metabolism, tumor migration, stress and inflammatory reaction. Immunocytochemistry and immunohistochemistry further confirmed these proteins present in the solid tumors. CONCLUSION: Distinct differential proteomics profiles exist in glioblastoma cell lines and normal glia cells, likely related to the transformation of normal glia to glioma.