Solanum Muricatum Promotes Osteogenic Differentiation of Rat Bone Marrow Stromal Cells

Solanum muricatum (SM), also known as pepino, is known for its antioxidative and anti‐inflammatory effects. The aim of this study was to evaluate the effects of SM extract in promoting osteogenic differentiation and regulating the Wnt and bone morphogenetic protein (BMP) signaling pathways. Ingredients of pepino were extracted and identified. SM extracts were used to treat rat bone marrow stromal cells (BMSCs), followed by evaluating alkaline phosphatase activities and mineralization levels. The mRNA levels of osteogenic biomarkers, including OPN and Collagen I, were also evaluated with real‐time polymerase chain reaction. After treatment with SM extracts, the expressions of key proteins in the Wnt and BMP signaling pathways were assessed. DKK‐1 and noggin, which are Wnt and BMP inhibitors, respectively, were added with SM extracts to investigate the role of Wnt and BMP pathways in the ameliorating effects of SM extract in osteogenesis. Treatment of BMSCs with SM extract promoted osteogenesis. Meanwhile, upregulations in the Wnt and BMP pathways were also observed. However, inhibiting both pathways compromised the effects of SM extract in promoting osteogenic differentiation. SM extract promotes osteogenic differentiation in BMSCs via promoting the Wnt and BMP signaling pathways.     

Protective effects of an aqueous extract from pepino (Solanum muricatum Ait.) in diabetic mice

BACKGROUND: This study analysed the content of ascorbic acid, phenolic acids and flavonoids in aqueous and ethanol extracts of pepino (Solanum muricatum Ait.), and examined the protective effects of pepino aqueous extract (PAE) in a mouse model of diabetes. PAE at 1, 2 and 4% was supplied for 5 weeks. RESULTS: Aqueous and ethanol extracts had similar levels of total phenolic acids, but PAE had a higher content of ascorbic acid and total flavonoids than the ethanol extract. PAE treatments at 2% and 4% significantly lowered plasma glucose level (P < 0.05); however, only the 4% PAE significantly elevated plasma insulin level at week 5 (P < 0.05). PAE treatments significantly decreased the levels of malonyldialdehyde and reactive oxygen species in kidney (P < 0.05); however, only the 2% and 4% treatments significantly reduced oxidised glutathione formation, increased glutathione level, and retained renal glutathione peroxidase and catalase activities (P < 0.05). PAE treatments at 2% and 4% significantly lowered renal interleukin (IL)‐6 and tumour necrosis factor‐α levels (P < 0.05); however, only the 4% treatments significantly diminished renal IL‐1β and levels of monocyte chemoattractant protein‐1 (P < 0.05). PAE treatments at 4% significantly decreased aldose reductase activity and sorbitol production in kidney (P < 0.05). CONCLUSION: These findings support the suggestion that pepino aqueous extract could attenuate the progression of diabetes via its antioxidative, anti‐inflammatory and antiglycative effects. Copyright © 2011 Society of Chemical Industry     

Effects of amla extract and collagen peptide on UVB-induced photoaging in hairless mice

Amla (Emblica officinalis Gaertn.) is a rich dietary source of vitamin C, minerals and amino acids, and also contains a wide variety of phenolic compounds. Amla has also been used as a principal constituent of many preparations of Ayurved, the Indian system of traditional medicine. Gelatin hydrolysate, also known as collagen peptide as a functional ingredient, is obtained from animal hide or fish scales. Ingestion of gelatin or collagen peptide affects various functions of the body, including bone, the achilles tendon, and skin. However, there are few data on the effects of amla extract and collagen peptide on photoaging in vivo. In the present study, therefore, we administered amla extract and/or collagen peptide to hairless mice that were repeatedly exposed of UVB irradiation, and examined the resulting effects on photoaging. Amla extract and collagen peptide suppressed the formation of 8-OHdG-positive cells and epidermal hyperplasia, and controlled skin hydration, thus reducing skin wrinkle formation in the mice. Collagen peptide, but not amla extract, also enhanced the production of collagen. We demonstrated that amla extract and collagen peptide exerted an additive effect in ameliorating skin dehydration and wrinkle formation, suggesting that they were able to attenuate photoaging effectively in UVB-irradiated hairless mice.     

Changes in degradation and phosphorylation level of titin in three ovine muscles during postmortem

The objective of this study was to investigate the degradation of titin and its phosphorylation level in three muscles from sheep. The MFI and pH from the longissimus lumborum (LL), semimembranosus (SM) and psoas major (PM) muscles were measured at 30 min, 1, 2, 7, 14, 21 and 28 days postmortem. Myofibrillar proteins were extracted, separated by SDS‐PAGE and quantified by phosphor‐specific staining. Phosphorylation of titin was predicted by Pro‐Q Diamond‐SYPRO Ruby staining. Two days after exsanguination, the pH and MFI of the PM were higher than those of the LL and SM muscles (P < 0.05). The sarcomere length of the PM muscle was also longer than that of the LL and SM muscle (P < 0.05). PM muscles had a highest phosphorylation level (P < 0.05) at 0.5 h postmortem and showed the greatest degree of titin degradation over 28 days. This suggests that phosphorylation of titin might accelerate its degradation.     

Inhibitory effect of Aspergillus niger on the growth of Aspergillus ochraceus and Aspergillus flavus,and on aflatoxin formation

Aspergillus ochraceus and A flavus were grown on synthetic media (SM) supplemented with 50 or 200 ml litre^?1 SM on which A niger had been grown previously ( ‘A niger medium’ = ANM). Controls included SM acidified to pH 6.0 or 4.4, SM diluted with 50 or 200 ml litre^?1 water, and diluted-acidified SM. For both fungi, higher growth inhibition was recorded on ANM-containing SM than in the controls. Aflatoxin formation was markedly inhibited on SM to which 20 ml litre^?1 ANM extract (in methanol/chloroform, 2:1 v) had been added, although the growth of A flavus on that medium was almost the same as that in the control. It is concluded that the inhibitory effect of A niger on the growth of fungi should not be attributed merely to pH reduction, but also, mainly, to metabolites produced by the fungus in the growth medium, even at early stages of its growth.     

产胶原酶的琥珀葡萄球菌分离鉴定及其培养优化研究

针对胶原蛋白难消化利用的现状,自烟台近海分离筛选产胶原蛋白降解酶的微生物,获得高产菌株SM12,并经形态观察、生理生化试验和16S rDNA鉴定,确定菌株SM12为琥珀葡萄球菌（Staphylococcus succinus）。通过单因素试验确定最佳培养基成分及发酵条件为20 g/L葡萄糖,15 g/L牛肉膏,20 g/L NaCl,10 g/L明胶,初始pH 7.5,培养温度37 ℃,接种量7.5%（V/V）,装液量100 mL/500 mL。在此最佳条件下,筛选的菌株SM12在发酵24 h后获得最大胶原酶活力185.32 U/mL,具备在水产加工下脚料高值化加工领域的应用潜力。     

MILK BASIC PROTEIN ENHANCES THE BONE STRENGTH IN OVARIECTOMIZED RATS

We studied the effects of milk basic protein (MBP) on bone metabolism in ovariectomized (OVX) rats. Five‐week‐old female Sprague‐Dawley rats were ovariectomized and fed a low‐calcium diet (0.009% Ca) for 5 weeks. The OVX rats were divided into three experimental groups: Control group (20% casein), MBP‐L group (19.9% casein, 0.1% MBP), and MBP‐H (19% casein, 1% MBP) of six animals. The rats were fed each experimental diet for 3 weeks. The bone breaking strength and energy of femur in the MBP‐H group were significantly higher than those in the control group. The bone breaking energy of femur in the MBP‐L group was also significantly higher than those in the control group. There were no differences in the amount of femoral calcium and phosphorus among the three groups, however, the amounts of femoral proline, hydroxyproline and hydroxylysine (typical amino acids of collagen) in the MBP groups were significantly higher than those in the control group. These data indicate that MBP in the whey protein increases the amount of the bone collagen and enhances the bone strength.     

Characterisation of Lactobacillus fermentum SM-7 isolated from koumiss, a potential probiotic bacterium with cholesterol-lowering effects

BACKGROUND: Lactic acid bacteria (LAB) have been demonstrated to have cholesterol‐reducing effects in many studies. RESULTS: Lactobacillus fermentum SM‐7 screened from ten LAB strains isolated from koumiss, a fermented milk drink, reduced cholesterol by 66.8%. It also showed acid and bile tolerance as well as antimicrobial activity against pathogenic Escherchia coli and Staphylococcus aureus. Lactobacillus fermentum SM‐7 cells assimilated 61.5% and co‐precipitated and absorbed 38.5% of the cholesterol in the media. Co‐precipitation of cholesterol with cholic acid increased rapidly at pH levels below 6. In vivo experiments using L. fermentum SM‐7 on artificially induced hyperlipidaemial ICR mice significantly decreased serum total cholesterol and total triglyceride levels, low‐density lipoprotein cholesterol concentrations and atherogenic index (P < 0.01), while serum high‐density lipoprotein cholesterol concentrations did not increase significantly (P > 0.05). The body weight and liver weight/body weight ratio of SM‐7 groups were lower than those of mice on a high‐cholesterol diet that were not given lactobacilli. There was no bacterial translocation in the liver, spleen or kidney of experimental mice. CONCLUSION: The results suggested that L. fermentum SM‐7 is a potential probiotic bacterium with cholesterol‐lowering effects. Copyright © 2010 Society of Chemical Industry     

Antihypertensive properties of aqueous extracts of vegetable leaf‐fortified bread after oral administration to spontaneously hypertensive rats

This study investigated the potential cardiovascular health benefits of leavened bread produced from wheat flour that contained 1%, 2% and 3% additions of leafy vegetable powders obtained from Amaranthus viridis (AO), Solanum macrocarpon (SM) or Telfairia occidentalis (TO). Dried breads were extracted with water at 60 °C followed by analysis for total polyphenolic content (TPC), as well as in vitro inhibitions of angiotensin‐converting enzyme and renin activities. HPLC analysis of the bread extracts indicated the presence of mainly rutin, gallic acid, myricetin and caffeic acid. TPC of the vegetable‐fortified breads was significantly (P < 0.05) higher (5.8–7.6 mg gallic acid equivalent, GAE/g) than that of control bread (5.5 mg GAE/g). Oral administration of 100 mg dried extract/kg body weight to spontaneously hypertensive rats led to reductions (up to 42 mmHg) in systolic, diastolic and mean arterial blood pressure in comparison with 20 mmHg for the control bread.     

Zinc deprivation inhibits extracellular matrix calcification through decreased synthesis of matrix proteins in osteoblasts

Scope: Zinc is implicated as an activator for bone formation, however, its influence on bone calcification has not been reported. This study examined how zinc regulates the bone matrix calcification in osteoblasts. Methods and Results: Two osteoblastic MC3T3‐E1 cell subclones (SC 4 and SC 24 as high and low osteogenic differentiation, respectively) were cultured in normal osteogenic (OSM), Zinc deficient (Zn–, 1 μM), or adequate (Zn+, 15 μM) media up to 20 days. Cells (SC 4) were also supplemented with (50 μg/mL) or no ascorbic acid (AA) in combination with Zinc treatment. Zn– decreased collagen synthesis and matrix accumulation. Although AA is essential for collagen formation, its supplementation could not compensate for Zinc deficiency‐induced detrimental effects on extracellular matrix mineralization. Zn– also decreased the medium and cell layer alkaline phosphatase ALP activity. This decreased ALP activity might cause the decrease of Pi accumulation in response to Zn–, as measured by von Kossa staining. Ca deposition in cell layers, measured by Alizarin red S staining, was also decreased by Zn^–. Conclusion: Our findings suggest that zinc deprivation inhibits extracellular matrix calcification in osteoblasts by decreasing the synthesis and activity of matrix proteins, type I collagen and ALP, and decreasing Ca and Pi accumulation. Therefore zinc deficiency can be considered as risk factor for poor extracellular matrix calcification.     

Core‐Shell Collagen Peptide Chelated Calcium/Calcium Alginate Nanoparticles from Fish Scales for Calcium Supplementation

We report simple methods for preparing collagen peptide chelated calcium (cpcc) and a novel cpcc‐loaded nanoparticle from marine fish scales for calcium supplementation. Cpcc nanoparticles have an average diameter of approximately 150 nm and a calcium content of up to 130.4 g/kg. Calcium alginate was selected to encapsulate cpcc for the preparation of core‐shell cpcc/calcium alginate nanoparticles. The core‐shell nanoparticles were mainly 200 to 500 nm in diameter. The ratio of calcium to sulfur was approximately 1.6:1. In vivo experiments indicated both cpcc and core‐shell cpcc were able to improve calcium absorption and prevent calcium deficiency. Especially core‐shell cpcc worked well to increase femur bone mineral density and femur calcium content in rats significantly. The study demonstrated that cpcc and core‐shell cpcc nanoparticles were ideal for calcium supplementation.     

Application of whey protein isolate in bone regeneration: Effects on growth and osteogenic differentiation of bone-forming cells

Recently, milk-derived proteins have attracted attention for applications in the biomedical field such as tissue regeneration. Whey protein isolate (WPI), especially its main component β-lactoglobulin, can modulate immunity and acts as an antioxidant, antitumor, antiviral, and antibacterial agent. There are very few reports of the application of WPI in tissue engineering, especially in bone tissue engineering. In this study, we tested the influence of different concentrations of WPI on behavior of human osteoblast-like Saos-2 cells, human adipose tissue-derived stem cells (ASC), and human neonatal dermal fibroblasts (FIB). The positive effect on growth was apparent for Saos-2 cells and FIB but not for ASC. However, the expression of markers characteristic for early osteogenic cell differentiation [type-I collagen (COL1) and alkaline phosphatase (ALP)] as well as ALP activity, increased dose-dependently in ASC. Importantly, Saos-2 cells were able to deposit calcium in the presence of WPI, even in a proliferation medium without other supplements that support osteogenic cell differentiation. The results indicate that, depending on the cell type, WPI can act as an enhancer of cell proliferation and osteogenic differentiation. Therefore, enrichment of biomaterials for bone regeneration with WPI seems a promising approach, especially due to the low cost of WPI.     

POSTMORTEM CHANGES IN MYOFIBRILLAR PROTEINS OF GOAT SKELETAL MUSCLES

Postmortem changes at 5C in myofibrillar proteins of longissimus dorsi (LD), biceps femoris (BF), semimembranosus (SM) and semitendinosus (ST) muscles and myofibrillar structure of LD muscle of goat were investigated. Muscle samples were immediately collected after killing, and from carcasses stored at 5C for 3, 6, 9, 12 and 20 days. The sodium dodecyl sulfate‐polyacrylamide gel electrophoresis of myofibrils indicated the appearance of a 30 kDa component, depending on the type of the muscles. A new 55 kDa component appeared in BF and SM muscles during postmortem. Titin I and nebulin also disappeared during storage. The disappearance of titin 1 and nebulin and the appearance of a 30 kDa component were confirmed by Western blot analysis. The Transmission Electron Microscopy studies showed that after 3 days postmortem, Z‐disks stayed unaltered. After 6 days postmortem, a little ultrastructural alteration was observed, and at 12 days postmortem a considerable degradation of Z‐disk ultrastructure was shown. The Z‐disk degradation, which results in the fragmentation of myofibrils and the appearance of 30 kDa components, is the major change observed in goat skeletal muscles during postmortem.     

A CONVENIENT DOMINANT SELECTION MARKER FOR GENE TRANSFER IN INDUSTRIAL STRAINS OF SACCHAROMYCES YEAST: SMRI ENCODED RESISTANCE TO THE HERBICIDE SULFOMETURON METHYL

The SMR1-410 gene of S. cerevisiae, encoding resistance to the herbicide sulfometuron methyl (SM), was used as a dominant selection marker in yeast replicating and yeast integrating vectors for the transformation of wild type strains of baking, brewing (ale and lager), distilling, wine and sake Saccharomyces yeasts. Transformation of lithium treated cells by a YEp vector resulted in transformation frequencies ranging from 200 to 8,000 transformants per 10 ug of DNA. Utilizing a yeast integrating vector with SMR1–410 as the only yeast DNA sequences, it was demonstrated that a single copy of SMR1–410 is sufficient to confer stably inherited SM resistance. Thus the SMR1–410 sequence has the unique ability to act as a selectable marker and to also provide a site for chromosomal integration. Since transformants were resistant to levels at least seven fold higher than wild type strains the resistance phenotype was clearly expressed and easily scored in all industrial strains tested. Unlike other selection markers derived from mammalian or bacterial cells, SMR1–410 is derived from S. cerevisiae. Thus industrial utilization of this marker as a means of genetically improving food and beverage strains of Saccharomyces yeasts by recombinant DNA technology is enhanced, as government regulatory agencies are likely to view it in a more favourable light.     

基于高效液相色谱-高分辨质谱法的非靶标代谢组学方法分析鉴别不同产地番茄

目的 分析不同产地来源番茄内源成分的差异及建立产地来源判别方法。方法 采用液相色谱-高分辨质谱（liquid chromatography - high resolution mass spectrometry,HPLC-HRMS）分析来源于全国5个地区105个番茄样品的乙醇-水提取物化合物组成,结合主成分分析(principal component analysis PCA)、正交偏最小二乘法判别分析(orthogonal partial least squares analysis discriminant analysis OPLS-DA)、双向正交偏最小二乘法判别分析(two-way orthogonal PLS-DA,O2PLS-DA)方法,对采集的HPLC-HRMS数据进行标志成分筛选和产地溯源模型构建。结果 以全部875个化合物为变量构建O2PLS-DA模型,筛选出其中99个对模型判别贡献较大的特征成分,重新构建了番茄产地的OPLS-DA判别模型,判别正确率达到99.05%。对特征成分进行了化合物结构分析,确定了其中18个有机酸、黄酮类特征化合物。结论 本研究建立的基于液相色谱-高分辨质谱的非靶标代谢组学方法能够科学、准确地用于番茄产地溯源,该研究为番茄产地溯源提供了一种新的策略。     

三文鱼骨胶原低聚肽钙增加SD大鼠的骨密度

以三文鱼骨为原料制备三文鱼骨胶原低聚肽,与钙离子螯合为三文鱼骨胶原低聚肽钙并作为受试物,以低中高剂量作用SD大鼠,并设置空白组和碳酸钙阳性对照组,检测体重、钙吸收率与储留率、股骨干重、股骨长、骨密度等重要指标,评价三文鱼骨胶原低聚肽钙的增加骨密度功能。结果显示,螯合物的总蛋白含量为65.37%,螯合率为52.56%,螯合物得率为43.12%;各试验组体重无显著性差异(p>0.05),中、高剂量组和碳酸钙组的钙吸收率分别为39.18%、40.52%、38.38%,储留率分别为38.80%、39.27%、37.58%,显著高于空白对照组36.31%,35.91%(p<0.05),且高剂量组明显优于同等钙含量的碳酸钙组(p<0.05);另外高剂量组的股骨干重、股骨中点直径分别为0.57(g)和2.40(mm),股骨近端、远端和中点密度分别为0.19、0.22、0.17(g/cm2),均高于空白组(p<0.05),与碳酸钙组无显著性差异。表明三文鱼骨胶原低聚肽钙具有良好的促钙吸收效果和增加骨密度的功能,为相关新型功能性食品的开发利用奠定良好的理论基础。     

Probiotic lactic acid bacteria from Kung‐Som: isolation,screening, inhibition of pathogenic bacteria

Lactic acid bacteria (LAB) were isolated from Kung‐Som at various fermentation periods. Only ten strains, named D2SM22, D6SM3, D6SM24, D6SM26, D8SM21, D10SM5, D10SM11, D10SM16, D10SM20 and D16SM26 showed a survival rate of more than 50% under the simulated gastric juice. After being subjected to simulated gastric juice, four strains (D6SM3, D8SM21, D10SM16 and D10SM20) showed a survival rate of more than 50% in simulated small intestinal juices. Growth of strain D6SM3, D8SM21 and D10SM16 under micro‐aerobic and anaerobic conditions was not different. Tested pathogenic strains (Escherichia coli, Staphylococcus aureus, Bacillus cereus, Vibrio parahaemolyticus and Salmonella sp.) were inhibited by probiotic LAB. However, none of strains could produce bacteriocins. All strains were identified as Lactobacillus plantarum. No differences in pH, acidity, LAB count and liking scores between Kung‐Som produced with starter culture and conventional method were observed (P > 0.01).     

Comparison of the thermal characteristics of connective tissue in loose structured and normal structured porcine M. semimembranosus

The aim of the present study was to characterise the thermal properties of connective tissue in loose structured porcine SM muscles in comparison to normal looking SM muscles and to see whether the meat quality traits were related to the properties of connective tissue. Samples from the muscles with loose structure and light colour were selected by visual assessment and normal looking SM muscles were used as a control (n = 25 loose structured + 25 control). The loose structured muscles had lower ultimate pH (pH_u) than the control muscles. The onset and peak temperatures of thermal shrinkage of intramuscular connective tissue (T_o and T_p, respectively) in loose structured muscles were similar to those of control muscles when the full set of data (25 loose structured + 25 control) were analysed. When the T_o and T_p data from muscles the exhibiting ten lowest and ten highest pH_u values were analysed, the low pH_u muscles (all classified as loose structured) had lower T_o and T_p than the high pH_u muscles (all classified as control) (p < 0.05). Drip loss of loose structured SM muscles (11.1%) was dramatically higher than that of control muscles (3.9%). Collagen content and collagen solubility were similar in loose structured and control muscles. It seemed that changes in the properties of intramuscular connective tissue were more easier found in porcine SM muscles with low pH_u than in SM muscles with high pH_u.     

Preparation of hoki (Johnius belengerii) bone oligophosphopeptide with a high affinity to calcium by carnivorous intestine crude proteinase

In the present study, the skeletons discarded from industrial processing of hoki (Johnius belengerii) were digested by a heterogeneous enzyme extracted from the intestine of a carnivorous fish (also discarded from industrial processing), bluefin tuna (Thunnus thynnus), in order to utilize the bone in nutraceuticals with a high bioavailability of calcium. The tuna intestine crude enzyme (TICE) could effectively biodegrade the hoki bone matrices composed of collagen, non-collageneous proteins, carbohydrates and minerals. A fish bone phosphopeptide (FBP) containing 23.6% of phoshorus was isolated from the hoki bone hydrolysates degraded by TICE using HA affinity chromatography and gel permeation chromatography. After the FBP, with a molecular mass of 3.5 kDa, was interacted with calcium, 41.1 mg/l of soluble calcium were maintained at 20 mM phosphate buffer (pH 7.8) without the formation of insoluble calcium phosphate. The results provide evidence that the carnivorous fish intestine enzyme (TICE) could degrade the teleost (J. belengerii) bone, and the fish bone oligophosphopeptide prepared by the enzymatic degradation of the bone could be utilized as a nutraceutical with a potential calcium-binding activity.