Effect of lifelong sucrose consumption at human-relevant levels on food intake and body composition of C57BL/6N mice.

Introduction: Controversies surround the issue if chronic consumption of a high-sugar diet is detrimental to health or not. This study investigates whether lifelong consumption of a higher sucrose diet will induce overeating, and obesity, and cause metabolic dysfunctions such as hyperglycemia and dyslipidaemia in C57BL/6N mice, compared to a lower sucrose diet. Methods: Male C57BL/6N mice at 3 weeks of age were randomized into consuming a diet with 25 or 10% kcal from sucrose for the rest of their lives. Body weight, food and water intake, fasting blood glucose, insulin, and lipid levels were measured at regular intervals. At the end of the study, organs and tissues were collected and gene expression was measured. Results: There was no discernible difference in the impact on food intake, body composition, glucose and lipid homeostasis, liver triglyceride content, life expectancy, as well as gene expression related to intermediary metabolism between mice fed a diet with 10 vs. 25% kcal as sucrose over their lifespan. We also showed that switching from a 25% kcal diet to a 10% kcal diet at different life stages, or vice versa, did not appear to affect these outcomes of interest. Discussion: The results from our study suggest that lifelong consumption of a higher sugar diet generally did not induce overeating and obesity, disrupt carbohydrate metabolism and lipid homeostasis, and reduce life expectancy compared with a lower sugar diet. Our unorthodox findings disagreed with the popular belief that higher sugar consumption is detrimental to health, which should be confirmed in future studies.

Polysaccharides extracted from Phellinus linteus ameliorate high-fat high-fructose diet induced insulin resistance in mice.

Phellinus linteus polysaccharide (PLP) has hypoglycemic effects, but mechanisms remain unclear. Male C57BL/6 J mice were either fed a normal diet (CON) or a high-fat high-fructose diet (HFD) for 16 weeks, and starting from week 12, HFD-fed animals in PLP group were orally given PLP. PLP administration significantly reduced fasting blood glucose level and ameliorated glucose intolerance. Differentially expressed genes involved in FOXO signaling pathway and in vitamin B12 (VB12) transport were identified between HFD and PLP group. HFD decreased the phosphatidylcholine (PC) to phosphatidylethanolamine (PE) ratio and S-adenosyl methionine to S-adenosyl homocysteine ratio, which were recovered by PLP treatment. Plasma VB12 levels in HFD group was lower than CON or PLP group, and PLP stimulated the proliferation of gut bacteria in genus Porphyromonas with capability of VB12 synthesis. In conclusion, PLP administration improved insulin resistance via modifying hepatic phospholipids metabolism and rescuing insulin signaling transduction.

Satisfaction, preference and error occurrence of three dry powder inhalers as assessed by a cohort naïve to inhaler operation.

Background: Inhaled medication is central to the treatment of COPD. Various types of inhaler devices, which directly deliver medication to the lung, have been developed. However, patients often exhibit incorrect techniques of inhaler usage. Effectiveness of therapy may be affected by the ease of device usage, size, convenience of use, durability, clarity of instructions and device preferences of patients. This study compares the satisfaction and preference, as well as error occurrence, with the use of Genuair®, Ellipta™ and Breezhaler™ by healthy subjects in Hong Kong. Subjects and methods: One hundred and thirty healthy Hong Kong Chinese subjects aged ≥40 years without a previous diagnosis of COPD and asthma and with no experience of using dry powder inhalers (DPIs) were recruited. Subjects learned to use the three DPIs by initially reading the instructions and then observing a demonstration with verbal explanation. The number of errors committed was evaluated. Subjects also completed a questionnaire to indicate their satisfaction and preference. Results: The satisfaction score of comfort for Breezhaler was significantly higher than that for Ellipta (p≤0.05), while the satisfaction score on confidence to have inhaled the entire dose was highest for Genuair compared with Ellipta (p≤0.0001) or Breezhaler (p≤0.05). The overall satisfaction score was significantly higher for Genuair than Ellipta (p≤0.05) or Breezhaler (p≤0.01). After reading the instructions, the highest number of subjects committing one or more critical errors was with Breezhaler (97) followed by Genuair (70) and then Ellipta (33). Demonstration reduced the number of critical errors made by subjects for each DPI to one third or lower. Conclusion: Breezhaler seemed to be more comfortable and easy to carry, but users made less critical errors when using Ellipta after reading the instructions only. Genuair provided the clearest indication of correct dose preparation and inhalation.

Comparative proteomic analysis of fibrotic liver of rats fed high fat diet contained lard versus corn oil.

INTRODUCTION: The influences of dietary fatty acids on the progress of chronic liver diseases have attracted lots of attentions, but the mechanisms of the effects of lipids rich in saturated fatty acids or PUFAs on hepatic fibrogenesis remain unclear. METHODS: Female Fischer 344 rats were fed normal chow or chow plus 20% (w/w) of corn oil or lard, respectively, and injected CCl4 twice a week for 4 weeks to induce liver fibrosis. Masson's staining was adopted to illustrate the fibrosis level. The mRNA expression level of α-SMA and the DNA methylation level of its promoter region were analyzed. A 2-DE gel based proteomic approach was constructed to investigate the differential expression level of hepatic proteome between three diet groups. RESULTS: Histological evaluations and α-SMA expression analysis illustrated the high corn oil intake has no effects on hepatic fibrogenesis, but lard intake aggravated liver fibrosis, partly attributed to DNA demethylation of α-SMA promoter region. 2-DE Gel based proteomic study demonstrated excessive lard consumption elevated the expression of fibrosis related alpha-1-antitrypsin precursor, and endoplasmic reticulum stress related proteins such as heat shock cognate 71 kDa, eukaryotic translation initiation factor 4A1 and protein disulfide isomerase associated 3. Moreover, unlike corn oil rich in PUFAs, lard had no effects to elevate the expression of glutathione S-transferases, but decreased the expression of iron store related proteins heme binding protein 1 and ferritin. CONCLUSIONS: Lard intake aggravates CCl4 induced liver fibrosis via enhancing the expression of fibrogenesis and ER stress related proteins, and disturbing the hepatic transmethylation reaction.

Glucagon-Like Peptide 2 Stimulates Postresection Intestinal Adaptation in Preterm Pigs by Affecting Proteins Related to Protein, Carbohydrate, and Sulphur Metabolism.

BACKGROUND: Exogenous glucagon-like peptide 2 (GLP-2) stimulates intestinal adaptation after resection in animal models of pediatric short bowel syndrome (SBS). It is unknown whether the molecular mechanisms of such GLP-2 effects are similar to those of postresection spontaneous adaptation. Using preterm pigs as a model, we hypothesized that GLP-2 treatment would change the intestinal proteome within the first week after resection, relative to individuals not resected or resected without GLP-2 treatment. MATERIALS AND METHODS: Two-day-old preterm pigs were subjected to resection of 50% distal small intestine and fed total parenteral nutrition without (SBS) or with GLP-2 infusion (3.5 µg/kg/h, SBS+GLP-2) for 5 days. The proteome of the remnant proximal intestine was compared among the SBS, SBS+GLP-2, and unresected pigs, through gel-based proteomics. RESULTS: Thirty-two proteins with differential expression were identified. Ten of these proteins were affected by the resection alone (ie, SBS vs unresected pigs). Five of these resection-responsive proteins and another 22 proteins were affected by GLP-2 infusion (ie, SBS+GLP-2 vs SBS or unresected pigs). Resection alone mainly affected cellular structural proteins, while the added GLP-2 treatment affected proteins involved in protein processing and the metabolism of protein, carbohydrate, and sulphur. CONCLUSION: In the first days following resection, proteins affected by resection plus GLP-2 treatment differed markedly from those affected by the spontaneous intestinal adaptation following resection alone. Whether more long-term GLP-2 treatment may affect the intestinal proteome following intestinal resection remains unknown.

Extra Virgin Olive Oil Reduced Polyunsaturated Fatty Acid and Cholesterol Oxidation in Rodent Liver: Is This Accounted for Hydroxytyrosol-Fatty Acid Conjugation?

The effects of extra virgin olive oil (EVOO) and carbon tetrachloride (CCl4) induced oxidative stress in rats were determined by the generation of isoprostanoids. These are known to be robust biomarkers to evaluate nonenzymatic and free radical related oxidation. Other oxidative stress biomarkers such as hydroxyeicosatetraenoic acid products (HETEs) and cholesterol oxidation products (COPs) were also determined. The rodents received a control diet, high-fat diet (20% w/w) composed of extra virgin olive oil (EVOO), corn oil (CO), or lard, and high-fat diets with CCl4 insult throughout the experimental period. The EVOO diet was found to suppress the formation of isoprostanoids and COPs compared to that of the control. EVOO also had a high total phenolic content and antioxidant activity compared to those of CO and lard and may be contributed to by the hydroxytyrosol component conjugated to fatty acids (HT-FA). This is the first study to identify HT-FA in EVOO, and it was 4-fold higher than that of olive oil, whereas none was found in corn oil. Furthermore, the EVOO diet showed reduced liver lipid vesicles in CCl4 treated rats compared to that of the control. However, liver toxicity measurements of AST (aspartate transaminase) and ALT (alanine transaminase) activities showed augmentation with CCl4 treatment but were not alleviated by the diets given. Our findings suggest that EVOO is a daily functional food capable of enhancing the antioxidant system for liver protection; the effect is potentially attributed to the phenolic and lipophenolic (phenol conjugated by fatty acids) content.

Differential protective effects of extra virgin olive oil and corn oil in liver injury: a proteomic study.

Extra virgin olive oil (EVOO) presents benefits against chronic liver injury induced by hepatotoxins such as carbon tetrachloride (CCl4); however, the protective mechanisms remain unclear. In the present study, a two-dimensional gel based proteomic approach was constructed to explore the mechanisms. Rats are injected with CCl4 twice a week for 4 weeks to induce liver fibrosis, and were fed laboratory chow plus 20% (w/w) of either corn oil or EVOO over the entire experimental period. Histological staining, MDA assay and fibrogenesis marker gene analysis illustrate that the CCl4-treated animals fed EVOO have a lower fibrosis and lipid peroxidation level in the liver than the corn oil fed group. The proteomic study indicates that the protein expression of thioredoxin domain-containing protein 12, peroxiredoxin-1, thiosulphate sulphurtransferase, calcium-binding protein 1, Annexin A2 and heat shock cognate 71 kDa protein are higher in livers from EVOO-fed rats with the CCl4 treatment compared with those from rats fed with corn oil, whereas the expression of COQ9, cAMP-dependent protein kinase type I-alpha regulatory subunit, phenylalanine hydroxylase and glycerate kinase are lower. Our findings confirmed the benefits of EVOO against chronic liver injury, which may be attributable to the antioxidant effects, hepatocellular function regulation and hepatic metabolism modification effects of EVOO.

Cordyceps cicadae induces G2/M cell cycle arrest in MHCC97H human hepatocellular carcinoma cells: a proteomic study.

BACKGROUND: Cordyceps cicadae is a medicinal fungus that is often used for treating cancer. However, the anticancer mechanisms of C. cicadae are largely unknown. This study aims to investigate the anticancer mechanisms of C. cicadae against hepatocellular carcinoma cells in vitro using a proteomic approach. METHODS: Human hepatocellular carcinoma MHCC97H cells were treated with a water extract of C. cicadae (0, 100, 250, 500, and 1000 µg/mL) for 48 h and harvested for cell viability assays. The significant differences in protein expression between control and C. cicadae-treated cells were analyzed by two-dimensional gel-based proteomics coupled with matrix-assisted laser desorption ionization-time of flight mass spectrometry. Flow cytometry analysis was employed to investigate the cell cycle and cell death. The anticancer molecular mechanism was analyzed by whole proteome mapping. RESULTS: The water extract of C. cicadae (0, 100, 250, 500, and 1000 µg/mL) inhibited the growth of MHCC97H cells in a dose-dependent manner via G2/M phase cell cycle arrest with no evidence of apoptosis. Among the identified proteins with upregulated expression were dynactin subunit 2, N-myc downstream-regulated gene 1, heat shock protein beta-1, alpha-enolase isoform 1, phosphatidylinositol transfer protein, and WD repeat-containing protein 1. Meanwhile, the proteins with downregulated expression were 14-3-3 gamma, BUB3, microtubule-associated protein RP/EB family member 1, thioredoxin-like protein, chloride intracellular channel protein 1, ectonucleoside triphosphate diphosphohydrolase 5, xaa-Pro dipeptidase, enoyl-CoA delta isomerase 1, protein-disulfide isomerase-related chaperone Erp29, hnRNP 2H9B, peroxiredoxin 1, WD-40 repeat protein, and serine/threonine kinase receptor-associated protein. CONCLUSION: The water extract of C. cicadae reduced the growth of human hepatocellular carcinoma MHCC97H cells via G2/M cell cycle arrest.

Individual and combined effects of Fusarium toxins on the mRNA expression of pro-inflammatory cytokines in swine jejunal epithelial cells.

Fusarium toxins have been arousing public interest in recent years because of their potential health hazards for humans and agricultural livestock. It was hypothesized that selected pro-inflammatory cytokines might serve as sensitive biomarkers of the predicted adverse effects of Fusarium toxins on the basis of their potential ability to induce immune and intestinal alterations comparable to those in human chronic inflammatory infection. Consequently, the aim of this study was to elucidate individual and combined effects of four common Fusarium toxins, deoxynivalenol (DON), nivalenol (NIV), zearalenone (ZEA) and fumonisin B1 (FB1) on the mRNA expression of pro-inflammatory cytokines (IL1α, IL1ß, IL6, IL8, TNFα and MCP-1) using a porcine jejunal epithelial cell line, IPEC-J2. Based on a dose-response relationship between individual mycotoxins and cell viability (MTT assay) that was previously established, cytotoxic and non-cytotoxic concentrations were selected to investigate combinations of two, three and all four of the mycotoxins. In general, up-regulation of pro-inflammatory cytokine mRNA expression occurred for both individual and mixtures of Fusarium toxins at cytotoxic concentrations, whereas significant up-regulation of pro-inflammatory cytokine mRNA mostly obtained when the toxins existed in mixtures at non-cytotoxic concentrations and these mixtures were found to cause cytotoxicity from MTT assay determined previously. Therefore, it may be concluded that some of the changes in the mRNA expression of IL1α, IL1ß, IL6, IL8, TNFα and MCP-1 could be cytotoxicity-related. It was also noted that additive effects were not always observed for the mixtures. These data suggest that individual or mixtures of Fusarium toxins could cause or exacerbate intestinal inflammation. These also provide a better understanding of the possible effects of Fusarium toxins, alone or in combinations on the immunological defense mechanisms of IECs, which would contribute to the risk assessment of these toxins.

Premature delivery reduces intestinal cytoskeleton, metabolism, and stress response proteins in newborn formula-fed pigs.

OBJECTIVE: Preterm infants often show intolerance to the first enteral feeds, and the structural and functional basis of this intolerance remains unclear. We hypothesized that preterm and term neonates show similar gut trophic responses to feeding but different expression of intestinal functional proteins, thus helping to explain why preterm neonates are more susceptible to feeding-induced disorders such as necrotizing enterocolitis (NEC). METHODS: Incidence of feeding-induced NEC, intestinal mass, and brush border enzyme activities, and the intestinal proteome in preterm cesarean-delivered pigs were compared with the corresponding values in pigs delivered spontaneously at term. RESULTS: For both preterm and term pigs, mucosal mass and maltase activity increased (50%-100%), whereas lactase decreased (-50%), relative to values at birth. Only preterm pigs were highly NEC sensitive (30% vs 0% in term pigs, P < 0.05). By gel-based proteomics, 36 identified proteins differed in expression, with most proteins showing downregulation in preterm pigs, including proteins related to intestinal structure and actin filaments, stress response, protein processing, and nutrient metabolism. CONCLUSIONS: Despite that enteral feeding induces rapid gut tropic response in both term and preterm neonates, the expression level of cellular proteins related to mucosal integrity, metabolism, and stress response differed markedly (including complement 3, prohibitin, ornithine carbamoyltransferase, and arginosuccinate synthetase). These proteins may play a role in the development of functional gut disorders and NEC in preterm neonates.

Intestinal proteome changes during infant necrotizing enterocolitis.

BACKGROUND: Changes in the intestinal and colonic proteome in patients with necrotizing enterocolitis (NEC) may help to characterize the disease pathology and identify new biomarkers and treatment targets for NEC. METHODS: Using gel-based proteomics, proteins in NEC-affected intestinal and colonic sections were compared with those in adjacent, near-normal tissue sections within the same patients. Western blot and immunohistochemistry were applied to crossvalidate proteomic data and histological location of some selected proteins. RESULTS: Thirty proteins were identified with differential expression between necrotic and vital small-intestine sections and 23 proteins were identified for colon sections. Five proteins were similarly affected in the small intestine and colon: histamine receptors (HRs), actins, globins, immunoglobulin, and antitrypsin. Two heat shock proteins (HSPs) were affected in the small intestine. Furthermore, proteins involved in antioxidation, angiogenesis, cytoskeleton formation, and metabolism were affected. Finally, secretory proteins such as antitrypsin, fatty-acid binding protein 5, and haptoglobin differed between NEC-affected and vital tissues. CONCLUSION: NEC progression affects different pathways in the small intestine and colon. HSPs may play an important role, especially in the small intestine. The identified secretory proteins should be investigated as possible circulating markers of NEC progression in different gut regions.

Antibiotics increase gut metabolism and antioxidant proteins and decrease acute phase response and necrotizing enterocolitis in preterm neonates.

BACKGROUND: The appropriate use of antibiotics for preterm infants, which are highly susceptible to develop necrotizing enterocolitis (NEC), is not clear. While antibiotic therapy is commonly used in neonates with NEC symptoms and sepsis, it remains unknown how antibiotics may affect the intestine and NEC sensitivity. We hypothesized that broad-spectrum antibiotics, given immediately after preterm birth, would reduce NEC sensitivity and support intestinal protective mechanisms. METHODOLOGY/PRINCIPAL FINDINGS: Preterm pigs were treated with antibiotics for 5 d (oral and systemic doses of gentamycin, ampicillin and metrodinazole; AB group) and compared with untreated pigs. Only the untreated pigs showed evidence of NEC lesions and reduced digestive function, as indicated by lowered villus height and activity of brush border enzymes. In addition, 53 intestinal and 22 plasma proteins differed in expression between AB and untreated pigs. AB treatment increased the abundance of intestinal proteins related to carbohydrate and protein metabolism, actin filaments, iron homeostasis and antioxidants. Further, heat shock proteins and the complement system were affected suggesting that all these proteins were involved in the colonization-dependent early onset of NEC. In plasma, acute phase proteins (haptoglobin, complement proteins) decreased, while albumin, cleaved C3, ficolin and transferrin increased. CONCLUSIONS/SIGNIFICANCE: Depressed bacterial colonization following AB treatment increases mucosal integrity and reduces bacteria-associated inflammatory responses in preterm neonates. The plasma proteins C3, ficolin, and transferrin are potential biomarkers of the colonization-dependent NEC progression in preterm neonates.

Protective effect of Phellinus linteus polysaccharide extracts against thioacetamide-induced liver fibrosis in rats: a proteomics analysis.

BACKGROUND: The hepatoprotective potential of Phellinus linteus polysaccharide (PLP) extracts has been described. However, the molecular mechanism of PLP for the inhibition of liver fibrosis is unclear. This study aims to investigate the molecular protein signatures involved in the hepatoprotective mechanisms of PLP via a proteomics approach using a thioacetamide (TAA)-induced liver fibrosis rat model. METHODS: Male Sprague-Dawley rats were divided into three groups of six as follows: Normal group; TAA group, in which rats received TAA only; and PLP group, in which rats received PLP and TAA. Liver fibrosis was induced in the rats by repeated intraperitoneal injections of TAA at a dose of 200 mg/kg body weight twice a week for 4 weeks. PLP was given orally at a dose of 50 mg/kg body weight twice a day from the beginning of the TAA treatment until the end of the experiment. The development of liver cirrhosis was verified by histological examination. Liver proteomes were established by two-dimensional gel electrophoresis. Proteins with significantly altered expression levels were identified by matrix-assisted laser desorption/ionization-time of flight/time of flight mass spectrometry and the differentially expressed proteins were validated by immunohistochemical staining and reverse transcription polymerase chain reaction. RESULTS: Histological staining showed a remarkable reduction in liver fibrosis in the rats with PLP treatment. A total of 13 differentially expressed proteins including actin, tubulin alpha-1C chain, preprohaptoglobin, hemopexin, galectin-5, glutathione S-transferase alpha-4 (GSTA4), branched chain keto acid dehydrogenase hterotetrameric E1 subunit alpha (BCKDHA), glutathione S-transferase mu (GSTmu); glyceraldehyde-3-phosphate dehydrogenase (GAPDH); thiosulfate sulfurtransferase (TFT); betaine-homocysteine S-methyltransferase 1 (BHMT1); quinoid dihydropteridine reductase (QDPR); ribonuclease UK114 were observed between the TAA and PLP groups. These proteins are involved in oxidative stress, heme and iron metabolism, cysteine metabolism, and branched-chain amino acid catabolism. CONCLUSION: The proteomics data indicate that P. linteus may be protective against TAA-induced liver fibrosis via regulation of oxidative stress pathways, heat shock pathways, and metabolic pathways for amino acids and nucleic acids.

Identification of liver proteins and their roles associated with carbon tetrachloride-induced hepatotoxicity.

Carbon tetrachloride (CCl(4)) is a common hepatotoxin used in experimental models to elicit liver injury. To identify the proteins involved in CCl(4)-induced hepatotoxicity, two-dimensional gel electrophoresis was employed followed by mass spectrometry - mass spectrometry (MS/MS) to study the differentially expressed proteins during CCl(4) exposure in the Fischer 344 rat liver proteome for 5 weeks. Ten spots with notable changes between the Control and CCl(4) groups were successfully identified. Among them, four proteins with significant up-regulation, namely calcium-binding protein 1, protein disulfide isomerase, mitochondrial aldehyde dehydrogenase precursor, and, glutathione-S-transferase mu1 and six proteins with significant down-regulation, namely catechol-O-methyltransferase, hemoglobin-alpha-2-chain, hemopexin precursor, methionine sulfoxide reductase A, catalase and carbonic anhydrase 3, were identified. The data indicates that CCl(4) causes hepatotoxicity by depleting oxygen radical scavengers in the hepatocytes. In this rat model, we profiled hepatic proteome alterations in response to CCl(4) intoxication. The findings should facilitate understanding of the mechanism of CCl(4)-induced liver injury.

Bacterial colonization affects the intestinal proteome of preterm pigs susceptible to necrotizing enterocolitis.

BACKGROUND: In newborns, colonizing bacteria and enteral nutrition are important for early gut development and immunity. However, in preterm newborns, bacterial colonization, coupled with enteral feeding, can lead to marked intestinal inflammation and disease such as necrotizing enterocolitis (NEC). We hypothesized that the initial bacterial colonization of the gut affects the intestinal proteome independently of enteral feeding. OBJECTIVE: To identify the intestinal proteins affected by the first colonizing bacteria by comparing the intestinal proteome in formula-fed preterm pigs reared under germ free (GF) or conventional conditions. METHODS: Gel-based proteomics of the small intestine to detect proteins that may play a part in the response of the immature intestine to bacterial colonization after birth. RESULTS: Fourteen proteins involved in stress response and detoxification (e.g. heat-shock proteins, peroxiredoxin 1), tissue metabolism and apoptosis (e.g. annexin 2), and some signal transduction pathways were differentially expressed between GF and conventionally reared pigs. CONCLUSION: The premature intestine is highly responsive to initial bacterial colonization and the specific bacteria-related proteome changes may contribute to the stress response that makes the immature intestine sensitive to the pro-inflammatory effects of enteral feeding.

Enteral feeding in utero induces marked intestinal structural and functional proteome changes in pig fetuses.

Intestinal adaptation from parenteral to enteral nutrition is crucial for survival and growth of newborns. Rapid feeding-induced gut maturation occurs immediately after birth in both preterm and term neonates, but it remains unclear whether the responses depend on factors related to birth transition (e.g. bacterial colonization, endocrine, and metabolic changes). We hypothesized that enteral feeding matures the immature intestine, even in fetuses before birth. Hence, control pig fetuses were compared with fetuses fed with milk formula for 24 h in utero. Gel-based proteomics showed that feeding-induced changes in 38 proteins, along with marked increases in intestinal mass and changes in activities of brush border enzymes. Physiological functions of the identified proteins were related to enterocyte apoptosis (e.g. caspase 1) and nutrient metabolism (e.g. citric acid cycle proteins). Many of the differentiated proteins were similar to those identified previously in preterm pigs fed with the same formula after birth, except that effects on proteins related to inflammatory lesions (e.g. heat shock proteins) were absent. Our results show that enteral feeding, independently of the birth transition, induces marked gut maturation and proteome change in the immature intestine. Hence, immediate postnatal feeding-induced gut changes are largely independent of factors related to the birth transition.

Regulatory properties of polysaccharopeptide derived from Coriolus versicolor and its combined effect with ciclosporin on the homeostasis of human lymphocytes.

OBJECTIVES: Lymphocyte homoeostasis is essential in inflammatory and autoimmune diseases. In search of natural fungal metabolites with effects on lymphocyte homoeostasis, we recently reported that polysaccharopeptide (PSP) from Coriolus versicolor exhibited ciclosporin-like activity in controlling aberrant lymphocyte activation. This object of this study was to investigate its effect on lymphocyte homoeostasis. This was done by investigating the mechanistic actions of PSP in relation to ciclosporin by performing cell cycle and cell death analysis of human lymphocytes in vitro. METHODS: We investigated the effect of PSP in the presence and absence of ciclosporin on cell proliferation, cell cycle, cell death, immunophenotype and cell cycle regulatory proteins in human lymphocytes. KEY FINDINGS: The data showed that PSP exhibited homoeostatic activity by promoting and inhibiting the proliferation of resting and phytohaemagglutinin (PHA)-stimulated lymphocytes, respectively. PHA-stimulated lymphocytes exhibited G0/G1 cell cycle arrest that was accompanied by a reduction of cyclin E expression with PSP treatment. Both PSP and ciclosporin blocked the reduction of the CD4/CD8 ratio in stimulated lymphocytes. PSP did not induce cell death in human lymphocytes, but the suppression of the Fasreceptor suggested a protective role of PSP against extrinsic cell death signals. These homoeostatic effects were more potent with combined PSP and ciclosporin treatment than with either fungal metabolite alone. CONCLUSIONS: Collectively, the results reveal certain novel effects of PSP in lymphocyte homoeostasis and suggest potential as a specific immunomodulatory adjuvant for clinical applications in the treatment of autoimmune diseases.

Polysaccharopeptides derived from Coriolus versicolor potentiate the S-phase specific cytotoxicity of Camptothecin (CPT) on human leukemia HL-60 cells.

BACKGROUND: Polysaccharopeptide (PSP) from Coriolus versicolor (Yunzhi) is used as a supplementary cancer treatment in Asia. The present study aims to investigate whether PSP pre-treatment can increase the response of the human leukemia HL-60 cells to apoptosis induction by Camptothecin (CPT). METHODS: We used bivariate bromodeoxyuridine/propidium iodide (BrdUrd/PI) flow cytometry analysis to measure the relative movement (RM) of the BrdUrd positively labeled cells and DNA synthesis time (Ts) on the HL-60 cell line. We used annexin V/PI flow cytometry analysis to quantify the viable, necrotic and apoptotic cells. The expression of cyclin E and cyclin B1 was determined with annexin V/PI flow cytometry and western blotting. Human peripheral blood mononuclear cells were used to test the cytotoxicity of PSP and CPT. RESULTS: PSP reduced cellular proliferation; inhibited cells progression through both S and G2 phase, reduced 3H-thymidine uptake and prolonged DNA synthesis time (Ts) in HL-60 cells. PSP-pretreated cells enhanced the cytotoxicity of CPT. The sensitivity of cells to the cytotoxic effects of CPT was seen to be the highest in the S-phase and to a small extent of the G2 phase of the cell cycle. On the other hand, no cell death (measured by annexin V/PI) was evident with the normal human peripheral blood mononuclear cells with treatment of either PSP or CPT. CONCLUSION: The present study shows that PSP increases the sensitization of the HL-60 cells to undergo effective apoptotic cell death induced by CPT. The pattern of sensitivity of cancer cells is similar to that of HL-60 cells. PSP rapidly arrests and/or kills cells in S-phase and did not interfere with the anticancer action of CPT. PSP is a potential adjuvant to treat human leukemia as rapidly proliferating tumors is characterized by a high proportion of S-phase cells.

The cell cycle effects of docosahexaenoic acid on human metastatic hepatocellular carcinoma proliferation.

Given the reported side effects associated with chemotherapy and surgical resection, dietary intervention with omega-3 polyunsaturated fatty acids (PUFAs) has been postulated to be an alterative way to prevent liver cancer progression and metastasis. We studied the effects of an omega-3 PUFA, docahexaenoic acid (DHA) on COX-2 expression and the cell cycle control machinery that co-ordinately regulate the HCC cells growth. Our data showed that DHA (0-200 microM) retarded proliferation of the human metastatic HCC cell line MHCC97L dose-dependently. In addition, inhibition of cyclin A/Cdk2 interfered with S-phase progression further in agreement with the result of bivariate flow cytometric analysis which indicated that DNA synthesis time (Ts) was significantly prolonged by DHA in MHCC97L. The N-myc oncogene, the heat shock proteins Hsp27 and glucose-related protein 78 (GRP78) as well as the antioxidant enzymes superoxide dismutase may play significant roles in the cell cycle control and reduced-proliferation of MHCC97L by DHA. Our data indicated that it is imperative to develop therapeutic strategy with omega-3 PUFA that simultaneously targets COX-2 and other cell cycle regulators in hepatocarcinogenesis. This study provides novel mechanistic insights into the modulation of DHA on human hepatocarcinoma.

Identification of Linkage Disequilibrium SNPs from a Kidney-Yang Deficiency Syndrome Pedigree.

In order to probe the genetic traits of Kidney-yang Deficiency Syndrome (KDS), we employed a national standard of KDS diagnosis for the collection of KDS subjects. Each candidate KDS subject from a typical family was diagnosed by 5 independent physicians of Traditional Chinese Medicine (TCM), and repeated for 3 years, all on the first Saturday of December. Fifteen samples of genomic DNA were isolated and genotyped by Affymetrix 100 K arrays of single nucleotide polymorphism (SNP). Then appropriate tools were used for the analysis of linkage disequilibrium (LD) and bioinformatic mining of LD SNPs. The results indicated that our procedure of TCM diagnosis can effectively collect KDS subjects and therefore provide substantial basis for the linkage analysis of KDS. Five SNPs (i.e. rs514207, rs1054020, rs7685923, rs10515889 and rs10516202) were identified as LD SNPs from this KDS family, representing an unprecedented set of LD SNPs derived from TCM syndrome. These SNPs demonstrate midrange linkage disequilibrium within the KDS family. Two genes with established functions were identified within 100 bp of these SNPs. One is Homo sapiens double cortin domain containing 5, which interacts selectively with mono-, di- or tri-saccharide carbohydrate and involves certain signaling cascades. Another one, leucyl-tRNA synthetase, is also a pleiotropic gene response to cysteinyl-tRNA aminoacylation and protein biosynthesis. In conclusion, KDS is involved in special SNP linkage disequilibrium in the intragenic level, and genes within the flanks of these SNPs suggest some essential symptoms of KDS. However, definitive evidence to confirm or exclude these loci and to establish their biological activities will be required.