Brazilian green propolis improves gut microbiota dysbiosis and protects against sarcopenic obesity.

INTRODUCTION: Brazilian green propolis is an important honeybee product that is considered beneficial for health. Here, we examined the therapeutic potential of dietary supplementation with propolis against sarcopenic obesity using Db/Db mice. METHODS: Db/m mice fed a normal diet alone and Db/Db mice fed normal diet alone, or supplemented with different amounts of propolis (0.08, 0.4 and 2%), were examined for effects on sarcopenic obesity. RESULTS: Propolis improved the glucose tolerance (P < 0.001), increased the grip strength (P < 0.001) and the weight of soleus (P = 0.006) and plantaris muscles (P = 0.008). Moreover, propolis improved the non-alcoholic fatty liver disease activity score (P < 0.001) and decreased the expression of genes related to inflammation, liver fibrosis and fatty acid metabolism. Propolis decreased the accumulation of saturated fatty acids in the liver and increased their excretion in faeces. With regard to the innate immunity, propolis decreased the ratio of M1 macrophages (P = 0.008) and Type 1 and 3 innate lymphoid cells to CD45-positive cells (P < 0.001) and increased the ratio of M2 macrophages (P = 0.002) and ILC2s (P = 0.007) in the liver. Additionally, propolis decreased the expression of genes related to muscle atrophy and inflammation and the concentration of saturated fatty acids in the soleus muscle. 16S rRNA phylogenetic sequencing revealed that propolis increased the Bacteroidetes/Firmicutes ratio, and the abundance of Butyricicoccus and Acetivibrio genera. Gut microbiota related to the pentose phosphatase pathway and glycerolipid metabolism was more prevalent after the administration of propolis. CONCLUSIONS: This is the first study to demonstrate that propolis can improve sarcopenic obesity by improving dysbiosis due to overeating and provides new insights into diet-microbiota interactions during sarcopenic obesity.

Extracellular lipidome change by an SGLT2 inhibitor, luseogliflozin, contributes to prevent skeletal muscle atrophy in db/db mice.

BACKGROUND: Diabetes mellitus increases the excretion of urinary glucose from the renal glomeruli due to elevated blood glucose levels. In the renal tubules, SGLT2 is expressed and reabsorbs the excreted urinary glucose. In the pathogenesis of diabetes mellitus, glucose reabsorption by SGLT2 is increased, and SGLT2 inhibitors improve hyperglycaemia by inhibiting this reabsorption. When urinary glucose excretion is enhanced, glucose supply to skeletal muscle may be insufficient and muscle protein catabolism may be accelerated. On the other hand, SGLT2 inhibitors not only ameliorate hyperglycaemia but also improve fatty acid metabolism in muscle, which may prevent muscle atrophy. METHODS: Eight-week-old male db/m mice or db/db mice were fed a standard diet with or without the SGLT2i luseogliflozin (0.01% w/w in chow) for 8 weeks. Mice were sacrificed at 16 weeks of age, and skeletal muscle and serum lipidomes, as well as skeletal muscle transcriptome, were analysed. RESULTS: Administration of SGLT2i led to not only decreased visceral fat accumulation (P = 0.004) but also increased soleus muscle weight (P = 0.010) and grip strength (P = 0.0001). The levels of saturated fatty acids, especially palmitic acid, decreased in both muscles (P = 0.017) and sera (P = 0.041) upon administration of SGLT2i, while the content of monosaturated fatty acids, especially oleic acid, increased in both muscle (P < 0.0001) and sera (P = 0.009). Finally, the accumulation of transcripts associated with fatty acid metabolism, such as Scd1, Fasn, and Elovl6, and of muscle atrophy-associated transcripts, such as Foxo1, Mstn, Trim63, and Fbxo32, decreased following SGLT2i administration. CONCLUSIONS: Intramuscular fatty acid metabolism and gene expression were influenced by the extracellular lipidome, which was modified by SGLT2i. Hence, secondary effects, other than the hypoglycaemic effects of SGLT2i, might lead to the alleviation of sarcopenia.

ILC2s Improve Glucose Metabolism Through the Control of Saturated Fatty Acid Absorption Within Visceral Fat.

Background and aims: Group 2 innate lymphoid cells (ILC2s) have been implicated in the regulation of metabolic homeostasis in mice. Methods: In this study, the role of ILC2s in white adipose tissue (WAT) was investigated using ST2, an IL-33 receptor that is expressed on ILC2 knockout mice. Results: The deficiency of ST2 decreased ILC2s in WAT, whereas ex-ILC2, which acquired group 1 innate lymphoid cell (ILC1)-like traits, was increased. This led to significant metabolic disorders such as visceral fat obesity, decreased browning in WAT, reduction of energy metabolism, and impaired glucose tolerance, compared to wild type (WT) mice. Those metabolic abnormalities of ST2-knockout (ST2KO) mice were not ameliorated by IL-33 administration, but impaired glucose tolerance and visceral fat obesity were significantly improved by transplantation of ILCs from the bone marrow of WT mice. The relative expression of Cd36 in WAT increased due to the deficiency of ST2, and the storage of saturated fatty acids in WAT of ST2KO mice was significantly higher than that of WT mice. Moreover, saturated fatty acids aggravated the chronic inflammation in adipocytes, promoted the differentiation of M1-like macrophages, and inhibited that of M2-like macrophages. Conclusions: Our results indicated that ILC2 regulates diet-induced obesity and chronic inflammation through the regulation of saturated fatty acid absorption in visceral adipose tissue.

Trans Fatty Acid Intake Induces Intestinal Inflammation and Impaired Glucose Tolerance.

Background and Aims: Many nutritional and epidemiological studies have shown that high consumption of trans fatty acids can cause several adverse effects on human health, including cardiovascular disease, diabetes, and cancer. In the present study, we investigated the effect of trans fatty acids on innate immunity in the gut by observing mice fed with a diet high in trans fatty acids, which have been reported to cause dysbiosis. Methods: We used C57BL6/J mice and fed them with normal diet (ND) or high-fat, high-sucrose diet (HFHSD) or high-trans fatty acid, high-sucrose diet (HTHSD) for 12 weeks. 16S rRNA gene sequencing was performed on the mice stool samples, in addition to flow cytometry, real-time PCR, and lipidomics analysis of the mice serum and liver samples. RAW264.7 cells were used for the in vitro studies. Results: Mice fed with HTHSD displayed significantly higher blood glucose levels and advanced fatty liver and intestinal inflammation, as compared to mice fed with HFHSD. Furthermore, compared to mice fed with HFHSD, mice fed with HTHSD displayed a significant elevation in the expression of CD36 in the small intestine, along with a reduction in the expression of IL-22. Furthermore, there was a significant increase in the populations of ILC1s and T-bet-positive ILC3s in the lamina propria in mice fed with HTHSD. Finally, the relative abundance of the family Desulfovibrionaceae, which belongs to the phylum Proteobacteria, was significantly higher in mice fed with HFHSD or HTHSD, than in mice fed with ND; between the HFHSD and HTHSD groups, the abundance was slightly higher in the HTHSD group. Conclusions: This study revealed that compared to saturated fatty acid intake, trans fatty acid intake significantly exacerbated metabolic diseases such as diabetes and fatty liver.

Group 3 Innate Lymphoid Cells Protect Steatohepatitis From High-Fat Diet Induced Toxicity.

Background and Aims: Emerging evidence has revealed that innate lymphoid cells (ILCs) play a key role in regulating metabolic disorders. Here, we investigated the role of group 3 ILCs (ILC3s) in the modulation of Non-alcoholic fatty liver disease (NAFLD). Methods: RORγ gfp/gfp (RORgt KI/KI) and Rag2-/- mice with the administration of A213, RORgt antagonist, fed with a high-fat-diet (HFD) for 12 weeks, were used. We performed flow cytometry, real time PCR, and lipidomics analysis of serum and liver, and used RAW264.7 cells and murine primary hepatocytes in vitro. Results: HFD increased ILC3s and M1 macrophages in the liver, and RORgt KI/KI mice deficient in ILC3 showed significant fatty liver, liver fibrosis and significantly increased palmitic acid levels in serum and liver. In addition, administration of A213 to Rag2-/- mice caused significant fatty liver, liver fibrosis, and a significant increase in serum and liver palmitate concentrations, as in RORgt KI/KI mice. Addition of palmitc acid stimulated IL-23 production in cell experiments using RAW264.7. IL-22 produced by ILC3s inhibited the palmitate-induced apoptosis of primary hepatocytes. Conclusions: HFD stimulates IL-23 production by M1 macrophages, thus promoting ILC3 proliferation, whereas IL-22 secreted by ILC3s contributes to the upregulation of hepatic lipid metabolism and has anti-apoptosis activity.

Analysis method for PCBs in reclaimed oil using a fast-GC triple stage quadrupole mass spectrometer with the 13-component quantitation method.

It is necessary for companies supplying reclaimed oil to analyze polychlorinated biphenyls (PCBs), because there is a possibility of the presence of contaminants due to trace-level PCBs in the reclaimed oil. However, common analysis methods of PCBs are time-consuming and complicated. Fast-GC triple stage quadrupole mass spectrometer with the 13-component quantitation method is an official method for analyzing PCBs in insulating oil in Japan. This method is extremely fast and simplified. The purpose of this study involves an investigation of the aforementioned fast and simple method for potential use in the analysis of reclaimed oil. Furthermore, it was attempted to combine the method with sample preparation involving only hexane dilution. The effect of sample dilutions corresponding to 100, 300, and 500 times was evaluated for reducing the matrix effect. The matrix effect was suppressed at a dilution ratio equal to or exceeding 300 times. Calibration curves of four points, namely 0.01, 0.05, 0.1, and 0.5 ng/mL, (ignored origin) by using an internal standard method were prepared for the 13 components. The square of regression coefficient (R2) values of all calibration curves exceeded 0.997. This method was adopted for the analysis of reclaimed oil containing 0.5 µg/mL PCBs, which corresponds to the judgment criteria, and accurate quantitation (accuracy value, 94.0-102%) and good repeatability (%RSD, 3.6%) were obtained. Furthermore, the required sensitivity was maintained even when 800 samples were analyzed without a cleaning ion source and an exchanging analysis column.

Spontaneous regression of diffuse intrahepatic recurrence with portal vein tumor thrombus after resection of hepatocellular carcinoma.

We report a rare case of spontaneous regression of diffuse intrahepatic recurrence with portal vein tumor thrombus (PVTT) after resection of hepatocellular carcinoma (HCC). A 68-year-old man with hepatitis C virus-related liver cirrhosis presented with a 40 mm tumor in the right anterior segment of the liver. The tumor was diagnosed as HCC by typical imaging findings and elevated serum alpha-fetoprotein (AFP) (716 ng/ml) and protein induced by vitamin K absence II (PIVKA II) (8,100 ng/ml). A right anterior sectionectomy of the liver was performed. Microscopically, the tumor was moderately differentiated HCC. Four months after resection, a computed tomography (CT) scan showed diffuse intrahepatic recurrence with PVTT. Serum AFP was 12,319 ng/ml and PIVKA II was 168,000 ng/ml. The patient did not receive any further treatment for HCC including herbal medicine, and stopped smoking. Two years and 5 months later, no lesion was detected on a CT scan when serum AFP was 1.9 ng/ml. Ischemia due to main portal vein occlusion and rapid tumor growth might have induced tumor regression in the present case. Moreover, abstention from smoking might have improved his immunological function.

[A case of jejunal adenocarcinoma with serum DUPAN-2 elevation].

A 50-year-old woman was admitted to our hospital because of abdominal pain and vomiting. Ileus with ulcerated jejunal tumor was diagnosed and biopsy revealed adenocarcinoma. Because her serum level of DUPAN-2 was high, she was examined by PET scan, which revealed that she had a left ovarian mass in addition to the jejunal tumor. Surgical resection was performed: both tumors were adenocarcinoma, but the ovarian tumor was considered to be metastatic clinically and histologically. Immunostaining for DUPAN-2 was positive in the both tumors. The serum level of DUPAN-2 returned to normal after the surgery, and has been within normal limits for about 3 years without any additional therapy. This case shows a possible relation between small bowel adenocarcinoma and DUPAN-2.

White ball appearance: predictor of effective variceal ligation in massive bleeding with an obscure bleeding point.

Although a purple-colored ball-like appearance (purple ball appearance) is typically observed on ligated varices during endoscopic variceal ligation (EVL), another endoscopic appearance of ligated varices (white ball appearance), which is observed after EVL at the bleeding site, have been reported. We encountered a case of massive variceal bleeding with an obscure bleeding point, where this appearance was useful in the confirmation of effective ligation. A 66-year-old man, who had liver cirrhosis with hepatocellular carcinoma, presented with hematemesis and melena. Although emergent endoscopy revealed a fibrin-plug on the esophageal varix, massive esophageal bleeding occurred and precluded direct visualization of the bleeding point during endoscopy. After multiple EVL, one ball-like elevation with a white color (white ball appearance) and multiple ball-like elevations with a red or purple color (purple ball appearance) were observed in the esophagus. Based on the presence of the white ball appearance, a predictor of effective ligation, we confirmed the ligation of the exact site of bleeding and complete cessation of bleeding. In fact, the ligated varix with the white ball appearance contained a ruptured point. This characteristic endoscopic appearance is useful for the assessment of effective ligation in massive variceal bleeding with an obscure bleeding point.

Preoperative evaluation of malignancy in intraductal papillary mucinous neoplasms of the pancreas, especially in relation to dysplastic epithelial changes.

BACKGROUND/AIMS: Intraductal papillary mucinous neoplasms have a better prognosis than ductal adenocarcinomas of the pancreas. The aim of this study was to evaluate the malignant potential of IPMNs by their preoperative images. METHODOLOGY: Forty-three intraductal papillary mucinous neoplasms were divided into 3 duct ectatic types using preoperative images (the main duct type, the branch duct type, and the mixed type), and into 2 groups using resected specimens (the malignant group including severe dysplasia based on the WHO classification and the benign group). The diameters of the tumor, main pancreatic duct and mural nodule were measured on the images. RESULTS: Two thirds of main duct type cases were in the malignant group. For the branch duct and mixed types, the diameters of the tumor and detectable mural nodules were larger in the malignant group than in the benign group. A tumor diameter larger than 3.5cm and a mural nodule diameter larger than 6mm were risk factors for malignancy (p < 0.05). CONCLUSIONS: The main duct type, a tumor larger than 3.5cm of the branch duct or mixed type, and a mural nodule larger than 6mm were all indicators of malignancy risk.

Treatment results of radiotherapy for carcinoma of the cervical esophagus.

The methods and results of treatment for cancer of the cervical esophagus differ from those for cancer of the thoracic esophagus. Our objective was to retrospectively review the outcome for cervical esophageal cancer patients treated with radiotherapy. Twenty-seven patients with carcinoma of the cervical esophagus treated with definitive radiotherapy from 1988 to 2002 were enrolled in the study. Clinical stage (UICC 1997) was stage I in five, II in six, III in 12 and IV in four. Concurrent head and neck malignancy was found in six patients (22%). The mean radiation dose was 66 Gy. Concurrent chemotherapy (cisplatin and 5-fluorouracil) was performed in 23 patients. The actuarial overall survival rates at 1, 3 and 5 years were 55.6%, 37.9% and 37.9%, respectively, with a median survival of 13.9 months. In the patients with stage I, the 3-year and 5-year survival rates were 75% and 75%, respectively. With univariate analysis, only two of the possible prognostic factors were found to actually influence survival: performance status (p < 0.01) and tumor length (p < 0.01). The survival of patients with cervical esophageal cancer remains poor. It is thought that organ preservation is possible by definitive chemoradiation for early cancer.

Treatment results of chemoradiotherapy for clinical stage I (T1N0M0) esophageal carcinoma.

PURPOSE: In 1991, we started a clinical prospective trial for operable esophageal carcinoma, foreseeing organ preservation, to assess the treatment results after definitive chemoradiotherapy (CRT) for clinical Stage I (T1N0M0) esophageal cancer. PATIENTS AND METHODS: Between 1992 and 2003, 63 patients were enrolled in this study. Tumor depth was mucosal cancer (T1a) in 23 and submucosal cancer (T1b) in 40. CRT consisted of 55-66 Gy/50-60 fractions (median, 59.4 Gy); from 1 to 3 cycles (median, 2) of concurrent chemotherapy (Cisplatin and 5-fluorouracil), followed by high-dose-rate intraluminal brachytherapy 10-12 Gy/2-3 fractions. RESULTS: The 5-year overall and cause-specific and disease-free survival rates were 66.4%, 76.3%, and 63.7%, respectively. The 5-year cause-specific survival rates for T1a and T1b cancer patients were 85.2% and 70.0%, respectively (p = 0.06). The 5-year disease-free survival rates for T1a and T1b were 84.4% and 50.5%, respectively (p < 0.01). Esophageal fistula as a late toxicity occurred in 2 patients (G4: 1; G5: 1), and esophageal stricture requiring a liquid diet occurred in 2 patients. Pericardial effusion was observed in 3 patients. CONCLUSION: We confirmed that patients with T1N0M0 esophageal carcinoma had their esophagus preserved in 89.2% of cases after definitive CRT, and the survival rates were equivalent to those of previous reports of surgery.

Appendix is a priming site in the development of ulcerative colitis.

AIM: The role of the appendix has been highlighted in the pathogenesis of ulcerative colitis (UC). The aims of this study were to elucidate the immuno-imbalances in the appendix of UC patients, and to clarify the role of the appendix in the development of UC. METHODS: Colonoscopic biopsy specimens of the appendix, transverse colon, and rectum were obtained from 86 patients with UC: active pancolitis (A-Pan; n = 15), active left-sided colitis (A-Lt; n = 25), A-Lt with appendiceal involvement (A-Lt/Ap; n = 10), inactive pancolitis (I-Pan; n = 14), and inactive left-sided colitis (I-Lt; n = 22), and from controls. In the isolated mucosal T cells, the CD4/CD8 ratio and proportion of activated CD4+ T cells were investigated, and compared with controls. RESULTS: In the appendix, the CD4/CD8 ratio significantly increased in A-Lt and A-Lt/Ap. The ratio in the appendix also tended to increase in A-Pan. In the rectum, the ratio significantly increased in all UC groups. In the appendix, the proportion of CD4+CD69+ (early activation antigen) T cells significantly increased in all UC groups. In the rectum, the proportion of CD4+CD69+ T cells significantly increased only in A-Pan. The proportion of CD4+HLA-DR+ (mature activation antigen) T cells significantly increased only in the rectum of A-Pan, but not in the other areas of any groups. CONCLUSION: The increased CD4/CD8 ratio and predominant infiltration of CD4+CD69+ T cells in the appendix suggest that the appendix is a priming site in the development of UC.

Handcrafted two-channel colonoscope for grasping-forceps-assisted resection of giant pedunculated polyps.

BACKGROUND: Colonoscopic polypectomy of giant pedunculated polyps has an increased risk of bleeding and is technically difficult. To facilitate the removal of the polyps, we handcrafted a two-channel colonoscope and applied it for grasping-forceps-assisted resection. METHODS: We easily handcrafted a two-channel colonoscope by taping a plastic tube along the shaft of a standard colonoscope and used it for the technique in 10 patients with 12 giant pedunculated polyps. OBSERVATIONS: The colonoscope with forceps assistance proved to be satisfactory for handling detachable and polypectomy snares. Immediate bleeding occurred in one patient because the detachable snare could not be maneuvered over the polyp. In 3 patients, the plastic tube became mobile during the procedure because the tape that attached the tube became loose. No other complications occurred. CONCLUSIONS: A handcrafted two-channel colonoscope for grasping-forceps-assisted resection of giant pedunculated polyps is effective for the prevention of postpolypectomy bleeding and the reduction of technical difficulties.

Open-biopsy-forceps technique for endoscopic removal of distally migrated and impacted biliary metallic stents.

BACKGROUND: Endoscopic removal of distally migrated and impacted biliary metallic stents is technically challenging. An open-biopsy-forceps technique for endoscopic removal of these migrated stents is described. METHODS: The technique was used in 4 patients with distally migrated and impacted covered metallic stents. A closed biopsy forceps was advanced through the stent mesh and opened within the stent to form an "anchor." With endoscope withdrawal, the stent was dislodged easily from the duodenum to the stomach. After grasping an end of the stent with a snare, the stent was removed by complete withdrawal of the endoscope. OBSERVATIONS: In all patients, the impacted stent was removed successfully. Mean time for removal was 10.2 minutes. Although ulceration was evident in the duodenal wall where the distal stent end was impacted in all patients, no other complication or adverse event was observed. CONCLUSIONS: The open-biopsy-forceps technique is useful for endoscopic removal of distally migrated and impacted biliary metallic stents.