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1.
Recombinant Escherichia coli, expressing the oleate hydratase gene of Stenotrophomonas maltophilia, was permeabilized by sequential treatments with 0.125 M NaCl and 2 mM EDTA. The optimal conditions for the production of 10-hydroxy-12,15(Z,Z)-octadecadienoic acid from α-linolenic acid by permeabilized cells were 35 °C and pH 7.0 with 0.1 % (v/v) Tween 40, 50 g permeabilized cells l?1, and 17.5 g α-linolenic acid l?1. Under these conditions, permeabilized cells produced 14.3 g 10-hydroxy-12,15(Z,Z)-octadecadienoic acid l?1 after 18 h, with a conversion yield of 82 % (g/g) and a volumetric productivity of 0.79 g l?1 h?1. These values were 17 and 168 % higher than those obtained by nonpermeabilized cells, respectively. The concentration, yield, and productivity of 10-hydroxy-12,15(Z,Z)-octadecadienoic acid obtained by permeabilized cells are the highest reported thus far.  相似文献   

2.
Growth factors are the key elements in wound healing signaling for cell migration, differentiation and proliferation. Platelet-rich plasma (PRP), one of the most studied sources of growth factors, has demonstrated to promote wound healing in vitro and in vivo. Adipose tissue is an alternative source of growth factors. Through a simple lipoaspirate method, adipose derived growth factor-rich preparation (adipose tissue extract; ATE) can be obtained. The authors set out to compare the effects of these two growth factor sources in cell proliferation and migration (scratch) assays of keratinocyte, fibroblast, endothelial and adipose derived stem cells. Growth factors involved in wound healing were measured: keratinocyte growth factor, epidermal growth factor, insulin-like growth factor, interleukin 6, platelet-derived growth factor beta, tumor necrosis factor alfa, transforming growth factor beta and vascular endothelial growth factor. PRP showed higher growth factor concentrations, except for keratinocyte growth factor, that was present in adipose tissue in greater quantities. This was reflected in vitro, where ATE significantly induced proliferation of keratinocytes at day 6 (p < 0.001), compared to plasma and control. Similarly, ATE-treated fibroblast and adipose stem cell cultures showed accelerated migration in scratch assays. Moreover, both sources showed accelerated keratinocyte migration. Adipose tissue preparation has an inductive effect in wound healing by proliferation and migration of cells involved in wound closure. Adipose tissue preparation appears to offer the distinct advantage of containing the adequate quantities of growth factors that induce cell activation, proliferation and migration, particularly in the early phase of wound healing.  相似文献   

3.
Spermatogonial stem cells (SSCs) are the only type of cells that transmit genes to the subsequent generations. The proliferation, cultivation and identification of SSCs in vitro are critical to understanding of male infertility, genetic resources and conservation of endangered species. To investigate the effects of glial cell-derived neurotrophic factor (GDNF) and leukemia inhibitory factor (LIF) on the proliferation of mouse SSCs in vitro, supplement of GDNF and/or LIF were designed to culture SSCs. The testes of 6–8 d mouse were harvested and digested by two-step enzyme digestion method. The SSCs and Sertoli cells were separated by differential plating. Then the SSCs were identified by alkaline phosphatase staining, RT-PCR and indirect immunofluorescence cell analysis. The cellular proliferation capacity was measured by methyl thiazolyl tetrazolium assay. The results showed that addition of 20 and 40 ng/ml of GDNF could strongly promote growth of mouse SSCs (p < 0.05). There was no significant difference between LIF treatment groups and the control group in promoting proliferation of the mouse SSCs (p > 0.05). However, the combination of 20 ng/ml GDNF and 1,000 U/ml LIF could significantly enhance the invitro proliferation of mouse SSCs (p < 0.05), and the OD490 value was 0.696 at day 5 of culture when the density of SSCs was 5–10 × 104 cells/ml.  相似文献   

4.
Thirty Chlorella and 30 Scenedesmus strains grown in nitrogen-stressed conditions (70 mg L?1 N) were analyzed for biomass accumulation, lipid productivity, protein, and fatty acid (FA) composition. Scenedesmus strains produced more biomass (4.02?±?0.73 g L?1) after 14 days in culture compared to Chlorella strains (2.57?±?0.12 g L?1). Protein content decreased and lipid content increased from days 8 to 14 with an increase in triacylglycerol (TAG) accumulation in most strains. By day 14, Scenedesmus strains generally had higher lipid productivity (53.5?±?3.7 mg lipid L?1 day?1) than Chlorella strains (35.1?±?2.8 mg lipid L?1 day?1) with the lipids consisting mainly of C16–18 TAGs. Scenedesmus strains generally had a more suitable FA profile with higher amounts of saturated fatty acids and monounsaturated fatty acids (MUFAs) and a smaller polyunsaturated fatty acid (PUFA) component. Chlorella strains had a larger PUFA component and smaller MUFA component. The general trend in the FA composition of Chlorella strains was oleic > palmitic > α-linolenic = linoleic > eicosenoic > heptadecenoic > stearic acid. For Scenedesmus strains, the general trend was oleic > palmitic > linoleic > α-linolenic > stearic > eicosenoic > palmitoleic > heptadecenoic acid. The most promising strains with the highest lipid productivity and most suitable FA profiles were Scenedesmus sp. MACC 401, Scenedesmus soli MACC 721, and Scenedesmus ecornis MACC 714. Although Chlorella sp. MACC 519 had lower lipid productivity, the FA profile was good with a lower PUFA component compared to the other Chlorella strains analyzed and a low linolenic acid concentration.  相似文献   

5.
The objective of this study was to determine if adenosine alters growth of mammary epithelium. Mouse mammary epithelial cells (NMuMG) were cultured in DMEM supplemented with 10% fetal calf serum. After serum starvation for 24h, EGF (0–100ng/ml) and/or adenosine (0–100μm ) was added. Adenosine at concentrations of 1, 10 or 100μm increased DNA synthesis significantly, when compared to control. Addition of epidermal growth factor (EGF) (10ng/ml) into 1 or 10μm adenosine showed the interaction in DNA synthesis between EGF and adenosine. A similar result was observed when 100μm adenosine added to various concentrations of EGF (0–100ng/ml). In the second mammary gland (thoracic) organ culture studies, mammary development scores were increased by adenosine (100μm ), EGF (100ng/ml) and adenosine plus EGF. These results indicate that the purine nucleoside adenosine stimulates mammary epithelial cell growth and interacts with EGF in DNA synthesis of mouse mammary epithelial cells.  相似文献   

6.
Mature seed samples of twenty-four Boraginaceae taxa collected from their natural habitats in Turkey were analysed by GC for total oil content and fatty acid composition. The range of total fat in the taxa varied between 7.0 and 35.7%. The amounts of palmitic (16:0) and stearic (18:0) acids determined were 5.65–17.81 and 1.49–5.08%, respectively. Mono-unsaturated fatty acids were in the range 8.83–55.32% for oleic, 0.22–6.21% for eicosenoic, 0.04–8.94% for erucic, and 0.08–2.71% for nervonic acid. Poly-unsaturated fatty acids were between 1.41 and 68.44% for linoleic, 0.12 and 43.0% for α-linolenic, 0.04 and 24.03% for γ-linolenic, and 0.02 and 14.59% for stearidonic acid. Total saturated (9.3–23.7%), mono-unsaturated (10.59–73.28%), and poly-unsaturated fatty acids (13.91–68.78%) varied substantially. Total unsaturated fatty acids ranged from 70.12 to 90.29%. There were significant differences between fatty acid profiles at taxa (P < 0.05) at genera levels, based on mono-unsaturated and poly-unsaturated fatty acid concentrations (P < 0.05). Segregation at the generic level by principle-component analysis was accomplished based on nine major fatty acids. The fatty acid patterns, their relative proportions, and quantities of unusual fatty acids as additional biochemical markers seem to be useful in the taxonomy of Boraginaceae at generic and infrageneric levels. All taxa are, in general, rich in linoleic and α-linolenic acids as essential fatty acids for dietary reference intakes. Seed oils of Symphytum, Anchusa, and Trachystemon orientalis for γ-linolenic acid and Echium for both γ-linolenic and stearidonic acid may be evaluated as alternative wild sources.  相似文献   

7.
A solventogenic strain of Clostridium sp. BOH3 produces extracellular α-amylase (7.15 U/mg protein) in reinforced clostridial medium supplemented with sugarcane bagasse hydrolysate (1 % w/v) and a small amount of starch (0.1 % w/v), which is essential for the expression of α-amylase. In the presence of α-amylase, BOH3 utilizes starch directly without any pretreatment and produces butanol almost equivalent (~90 %) to the production of butanol from glucose. α-Amylase can be purified from culture supernatant by using one-step weak anion exchange chromatography with a yield of 43 %. In peptide fingerprinting analysis, this enzyme shows homology with α-amylase produced by Clostridium acetobutylicum ATCC824. However, the molecular weight is 54 kDa, which is smaller than α-amylase of ATCC824 (84 kDa). This enzyme has optimum temperature at 45–50 °C and optimum pH at 4.5–5.5. Under this condition, the enzyme activity is 91.32 U/mg protein, and its K m and V max values are 1.71?±?0.02 mg/ml and 96.13?±?0.15 μmol/min/mg protein, respectively. Activity of this α-amylase can be enhanced (>1.5 times) by addition of Ca2+ and Co2+ and its activity can be maintained at an acidic pH (pH 3–5) for about 24 h. These unique characteristics suggest that this enzyme can be used for saccharification of starch for production of biofuel in one pot.  相似文献   

8.
Islet cell transplantation is a major treatment strategy for type I diabetes, and has proven to be effective for maintaining glucose homeostasis. However, this treatment requires an extended period of immunosuppression to prevent rejection and recurrent transplantation to maintain function. Thus, to enhance the properties of transplanted islet cells, we examined the effect of the co-culture of luteal cells, which secrete progesterone, on islet cell viability, functionality, and revascularization. It was found that islet viability and functionality were higher in the co-cultured group than in single cultures of islets at 48 and 96 h, in parallel with increased progesterone and vascular endothelial growth factor (VEGF) secretion from luteal cells. In the co-culture groups, VEGF levels at 48 and 96 h and CD31 levels at 48 h were significantly higher than those in the islet groups (p?<?0.001 and p?<?0.05, respectively), and basic fibroblast growth factor (bFGF) levels were increased at 96 h (p?<?0.001). Thus, co-culture with luteal cells may increase islet vascularity by enhancing VEGF and bFGF levels for up to 96 h, which could help to markedly increase the pre-transplantation time to allow for effective immunosuppression therapy. This method may also promote islet cell viability and functionality. Progesterone and angiogenic factors secreted from luteal cells may be responsible for these positive effects.  相似文献   

9.
The Helicobacter pylori outer membrane proteins play an important role in pathogenesis; the outer inflammatory protein A (OipA) is one of these proteins which play the main role in the development of inflammation. In this study, purification of recombinant H. pylori OipA was performed by Ni–NTA affinity chromatography. Gastric carcinoma epithelial cells (AGS cell) were treated by different concentrations of recombinant OipA for various lengths of time and cell viability was evaluated by the viability assay. Statistical analysis showed that OipA had toxic effects on AGS cells in a concentration of 500 ng/ml after 24 and 48 h, and this toxic dose was 256 ng/ml after 72 h. OipA had direct toxic effects on gastric epithelial cells and the toxicity was observed to depend on time and dose of H. pylori exposure. Attachment of H. pylori to gastric epithelial cells is a key part in the pathogenesis and enables H. pylori to damage the epithelial cells with OipA.  相似文献   

10.
Vibrio alginolyticus, Vibrio campbellii, and Vibrio harveyi were inhibited by Isochrysis galbana in batch cultures. I. galbana reduced the V. alginolyticus, V. campbellii, and V. harveyi counts to undetectable levels in 2, 4, and 7 days (<0.01 Vibrio spp. mL?1), respectively, remaining so until the end of the experiment on day 15. Other heterotrophic bacteria reached counts of 106 CFU mL?1 on ZoBell medium at the end of the experiment. Vibrio parahaemolyticus was not inhibited by I. galbana. In all mixed I. galbana and Vibrio spp. cultures, the algal density increased from 3.5 to 4.0?×?107 cells mL?1, higher than that in I. galbana cultures alone, indicating a lack of an inhibitory effect on microalgae in the mixed cultures. The predominant fatty acids (>82 %) of I. galbana during the stationary growth phase were estearidonic (24.3 %), oleic (15.7 %), myristic (13.8 %), docosahexaenoic (11.0 %), palmitic (10.3 %), and α-linolenic (7.2 %) acids. These results demonstrate that I. galbana synthesizes antibacterial fatty acids that inhibit the growth of pathogenic bacteria such as V. alginolyticus, V. campbellii, and V. harveyi.  相似文献   

11.
Breast epithelial cells develop into polarized and highly organized acinar and ductal structures in response to stromal cues, including extracellular matrix composition and density, which can in part be reproduced in 3D culture conditions. Here, we present the effects of various 3D in vitro stroma compositions (termed “matrices” or “substrates”) on the ability of heterotypic cultures of epithelial and mesenchymal stem cells to organize into acinar and tubular structures. Normal murine mammary gland (NMuMG) cells were cultured, either alone or in combination (30:70) with mouse mesenchymal stem cells (D1), in 3D matrices generated by agarose, collagen, and Matrigel® alone or by a combination thereof. After 3–5 d in culture, cell distribution, organization, and the presence of acinus-like and tubule-like structures were determined. The number of acinar structures was significantly higher in cultures grown in combination matrices of agarose with Matrigel® or collagen I when compared with cultures grown in Matrigel® or collagen I alone (p?p?相似文献   

12.
Cell death-inducing DNA fragmentation factor-alpha-like effector A (CIDE-A) was first identified by its sequence homology with the N-terminal domain of DNA fragmentation factor (DFF). CIDE-A negatively regulates the activity of uncoupling protein 1 (UCP1) in brown adipose tissue. CIDE-A and UCP1 mRNA were detected by RT-PCR in cloned bovine mammary epithelial cells (bMEC) and lactating bovine mammary glands. Physiological concentrations of saturated fatty acids (stearate and palmitate), but not unsaturated fatty acids (oleate and linoleate) induced up-regulation of CIDE-A mRNA in bMEC. Treatment with insulin (5-10 ng/ml) induced down-regulation of CIDE-A and UCP1. The expression levels of CIDE-A and UCP1 mRNA in bovine mammary glands at various stages of the lactation cycle were determined by quantitative RT-PCR analysis. CIDE-A mRNA expression at peak lactation (2 months after parturition) was significantly higher than at dry off and non-pregnancy but not late lactation. These results suggest that CIDE-A and UCP1 are regulated by insulin and/or fatty acids in mammary epithelial cells and lactating mammary glands, and thereby play an important role in lipid and energy metabolism.  相似文献   

13.
Epidermal growth factor, a 6,400-dalton polypeptide from the mouse submaxillary gland, binds specifically to cells and membranes derived from a variety of human, rat, mouse, and bovine tissues. Liver, placenta, skin, cornea, and cultured chondrocytes, Hela cells, and Chang liver cells bind large amounts of epidermal growth factor, whereas fat cells, resting and lectin-stimulated human peripheral lymphocytes, mouse thymocytes, cultured rat hepatoma cells, and mammary cells from virgin and pregnant mice bind little or no epidermal growth factor. The binding site for epidermal growth factor is distinct from receptors for other anabolic peptides such as insulin, nerve growth factor, and growth hormone. The binding of epidermal growth factor is rapid and reversible. The rate constant of association is approximately 106 mole?1 sec?1, the rate constant of dissociation is about 6 × 10?4 sec?1, and the apparent equilibrium dissociation constant is about 10?9m. Trypsin at low concentrations (50–200 μg/ml) destroys the receptor site for epidermal growth factor. The binding of epidermal growth factor by membranes is not accompanied by appreciable degradation of the peptide present in the medium or of that bound to the membranes. Use can be made of the high affinity and specificity of membranes for epidermal growth factor to measure by a competitive binding assay as little as 200 pg of EGF per ml (3 × 10?11m).  相似文献   

14.
15.
Monocyte differentiation antigen CD14 is considered an important cell-activating mediator of inflammatory responses that may result in atherosclerosis, coronary heart disease (CHD), thrombus formation, and myocardial infarction (MI). A common C-260T polymorphism in the promoter of the CD14 gene, the trans-membrane receptor of lipopolysaccharides, has been inconsistently associated with CHD. To investigate this inconsistency, we performed a meta-analysis of 28 studies involving a total of 13,335 CHD cases and 7,979 controls for C-260T of the CD14 gene to evaluate the effect of CD14 on genetic susceptibility for CHD. An overall random effects odds ratio of 1.24 (95 % CI: 1.12–1.36, P < 10?5) was found for T allele. Significant results were also observed using dominant (OR = 1.34, 95 % CI: 1.17–1.54, P < 10?4) or recessive genetic model (OR = 1.25, 95 % CI: 1.10–1.41, P = 0.0004). There was strong evidence of heterogeneity (P < 10?5), which largely disappeared after stratification by ethnicity. After stratified by ethnicity, significant results were found in East Asians; whereas no significant associations were found among Caucasians and other ethnic populations in all genetic models. In the stratified analysis according to sample size, CHD endpoints, and HWE status, significantly increased risks for the polymorphism were found in all genetic models. In conclusion, our results indicate that the CD14 C-260T polymorphism is a risk factor of CHD, especially in East Asians. However, additional very large-scale studies are warranted to confirm our results.  相似文献   

16.
Anaemia is a widespread problem especially in the tropics. Among adolescent girls, it has negative consequences on growth, school performance, morbidity and reproductive performance. A cross-sectional study was conducted to investigate the prevalence of anaemia, iron, folate, zinc and copper deficiencies amongst adolescent schoolgirls in New Halfa, eastern Sudan, and to examine the relationship of these micronutrients with haemoglobin (Hb) levels. Out of 187 adolescent schoolgirls, 181 (96.8%) had anaemia (Hb?<?12 g/dl); 21% had mild anaemia (Hb 11.0–11.9 g/dl); 66.8.1% had moderate anaemia (Hb 8.0–10.9 g/dl), and 12.1% had severe anaemia (Hb?<?8 g/dl), respectively. Iron deficiency (S-ferritin?<?12 μg/l), iron deficiency anaemia (<12 m/dl and S- ferritin?<?12 μg/l) and folate deficiency (S-folate?<?3 ng/ml) were prevalent in 17.6%, 16.5% and 69% of these girls, respectively. Nine percent and 5.9% of these girls had zinc (<75 μg/ml) and copper deficiency (<75 μg/ml), respectively. Twenty-six (14%) girls had ≥2 micronutrient deficiencies. S-ferritin and zinc were significantly lower in patients with severe anaemia. Haemoglobin levels were significantly positively correlated with zinc levels (r?=?0.161, P?=?0.03) and with copper levels (r?=?0.151, P?=?0.03). Thus, interventions are required to prevent and control anaemia in this setting. Further research is needed.  相似文献   

17.
Although α-linolenic acid is nearly absent from Cyanidium caldarium cultured at 53 °C, it is the most abundant unsaturated fatty acid in 20 °C-grown cells. A sudden growth temperature shift of 55 to 25 °C does not stimulate the immediate biosynthesis of α-linolenic acid. However, after an induction period of 48 h, synthesis of α-linolenic acid from acetate can be detected, and the fatty acid accumulates in phosphatidyl choline and sulfolipid. The newly synthesized α-linolenic acid appears to be formed primarily by de novo synthesis and to a much lesser extent from the elongation of a previously formed hexadecatrienoic acid precursor. On the other hand, when a cell-free algal preparation was presented with a hexadecatrienoic acid precursor in the presence of [14C] malonyl-CoA, the α-linolenic acid formed demonstrated a synthesis by elongation of the precursor. While the cell appears enzymatically capable of α-linolenic acid biosynthesis by both the de novo and elongation processes, de novo synthesis of α-linolenic acid appears to be the more significant mode of synthesis.  相似文献   

18.
Free fatty acids (FFAs) in plasma are essential substrates for de novo synthesis of milk fat, or directly import into mammary cells. The physico-chemical properties of mammary cells membrane composition affected by FFAs with different chain lengths and saturability are unclear yet. Employing GC, FTIR and fluorescence spectroscopy, the adsorption capacity, phospholipids content, membrane proteins conformation, lipid peroxidation product, and free sulfhydryl of plasma membranes (PMs) interacted with different FFAs were determined. The mammary cells PMs at 38 and 39.5 °C showed different adsorption capacities: acetic acid (Ac) > stearic acid (SA) > β-hydroxybutyric acid (BHBA) > trans10, cis12 CLA. In the FTIR spectrum, the major adsorption peaks appeared at 2920 and 2850 cm?1 for phospholipids, and at 1628 and 1560 cm?1 for membrane proteins. The intensities of PMs-FFAs complexes were varied with the FFAs species and their initial concentrations. The β-sheet and turn structures of membrane proteins were transferred into random coil and α-helix after BHBA, SA and trans10, cis12 CLA treatments compared with Ac treatment. The quenching effects on the fluorescence of endogenous membrane protein, 1, 8-ANS, NBD-PE, and DHPE entrapped in PMs by LCFA were different from those of short chain FFAs. These results indicate that the adsorption of FFAs could change membrane protein conformation and polarity of head group in phospholipids. This variation of the mammary cells PMs was regulated by carbon chain length and saturability of FFAs.  相似文献   

19.
Expression of the human granulocyte–macrophage colony-stimulating factor (hGM-CSF) gene under the control of the 5′-regulatory sequence of the goat alpha-S1-casein gene with and without a matrix attachment region (MAR) element from the Drosophila histone 1 gene was studied in four and eight transgenic mouse lines, respectively. Of the four transgenic lines carrying the transgene without MAR, three had correct tissues-specific expression of the hGM-CSF gene in the mammary gland only and no signs of cell mosaicism. The concentration of hGM-CSF in the milk of transgenic females varied from 1.9 to 14 μg/ml. One line presented hGM-CSF in the blood serum, indicating ectopic expression. The values of secretion of hGM-CSF in milk of 6 transgenic lines carrying the transgene with MAR varied from 0.05 to 0.7 μg/ml, and two of these did not express hGM-CSF. Three of the four examined animals from lines of this group showed ectopic expression of the hGM-CSF gene, as determined by RT-PCR and immunofluorescence analyses, as well as the presence of hGM-CSF in the blood serum. Mosaic expression of the hGM-CSF gene in mammary epithelial cells was specific to all examined transgenic mice carrying the transgene with MAR but was never observed in the transgenic mice without MAR. The mosaic expression was not dependent on transgene copy number. Thus, the expected “protective or enhancer effect” from the MAR element on the hGM-CSF gene expression was not observed.  相似文献   

20.
Therapeutic administration of mesenchymal stem cells (MSCs) by systemic delivery utilizes the innate ability of the cells to home to damaged tissues, but it can be an inefficient process due to a limited knowledge of cellular cues that regulate migration and homing. Our lab recently discovered that a potent pro-inflammatory cytokine, macrophage migration inhibitory factor (MIF), inhibits MSC migration. Because MIF may act on multiple cellular targets, an activating antibody (CD74Ab) was employed in this study to examine the effect of one MIF receptor, CD74 (major histocompatibility complex class II-associated invariant chain), on MSC motility. CD74 activation inhibits in a dose-dependent manner up to 90% of in vitro migration of MSCs at 40 μg/ml CD74Ab (p?<?0.001), with consistent effects observed among three MSC donor preparations. A blocking peptide from the C-terminus of CD74 eliminates the effect of CD74Ab on MSCs. This suggests that MIF may act on MSCs, at least in part, through CD74. Late-passage MSCs exhibit less chemokinesis than those at passage 2. However, MSCs remain responsive to CD74 activation during ex vivo expansion: MSC migration is inhibited ~2-fold in the presence of 5 µg/ml CD74Ab at passage 9 vs. ~3-fold at passage 2 (p?<?0.001). Consistent with this result, there were no significant differences in CD74 expression at all tested passages or after CD74Ab exposure. Targeting CD74 to regulate migration and homing potentially may be a useful strategy to improve the efficacy of a variety of MSC therapies, including those that require ex vivo expansion.  相似文献   

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