Preparation of Puerarin Long Circulating Liposomes and Its Effect on Osteoporosis in Castrated Rats.

Osteoporosis is a disease that causes low bone mass and deterioration of bone microarchitecture. Puerarin is a natural isoflavone compound that has been shown to possess anti-inflammatory, antioxidant and ameliorative effects on osteoporosis with less adverse reactions. However, its fast metabolism and low oral bioavailability limit its application. This study aimed to prepare D-α-tocopherol polyethylene glycol 1000 succinate (TPGS)- modified Puerarin Long Circulating Liposomes (TPGS-Puerarin-liposomes), in order to improve the oral bioavailability of puerarin, before evaluation of its pharmacological activity in vitro and in vivo. We employed film dispersion method to develop TPGS-Puerarin-liposomes before appropriate characterizations. Afterwards, we utilized in vivo imaging, pharmacokinetic analysis and in vitro drug release testing to further evaluate the in vivo and in vitro delivery efficiency. In addition, we established a castrated osteoporosis rat model to observe the changes in femur tissue structure and bone micromorphology via hematoxylin-eosin (HE) staining and Micro Computed Tomography (Micro CT). Besides, levels of oxidative stress and inflammatory indicators, as well as expression of wnt/ß-catenin pathway-related proteins were detected. In terms of physiochemical properties, the respective mean particle size (PS) and zeta potential (ZP) of TPGS-Puerarin-liposomes were 76.63±0.59 nm and -25.54±0.11 mV. The liposomal formulation exhibited encapsulation efficiency (EE) of 95.08±0.25% and drug loading (DL) of 7.84±0.07%, along with excellent storage stability. Compared with free drugs, the TPGS-Puerarin-liposomes demonstrated a sustained release effect and could increase blood concentration of puerarin in rats, thereby significantly improving its bioavailability. Also, in vivo studies have confirmed potential of the liposomes to promote bone tissue targeting and accumulation of puerarin, coupled with significant improvement of the osteoporotic status. Besides, the liposomes could also reduce levels of oxidative stress and inflammatory factors in serum and bone tissue. Additionally, we discovered that TPGS-Puerarin-liposomes increased Wnt, ß-catenin and T-cell factor (TCF) expressions at protein level in the wnt/ß-catenin signaling pathway. This study has demonstrated the potential of TPGS-Puerarin-liposomes for treatment of osteoporosis.

Unveiling the mechanism of efficient ß-phenylethyl alcohol conversion in wild-type Saccharomyces cerevisiae WY319 through multi-omics analysis.

ß-Phenylethanol (2-PE), as an important flavor component in wine, is widely used in the fields of flavor chemistry and food health. 2-PE can be sustainably produced through Saccharomyces cerevisiae. Although significant progress has been made in obtaining high-yield strains, as well as improving the synthesis pathways of 2-PE, there still lies a gap between these two fields to unpin. In this study, the macroscopic metabolic characteristics of high-yield and low-yield 2-PE strains were systematically compared and analyzed. The results indicated that the production potential of the high-yield strain might be contributed to the enhancement of respiratory metabolism and the high tolerance to 2-PE. Furthermore, this hypothesis was confirmed through comparative genomics. Meanwhile, transcriptome analysis at key specific growth rates revealed that the collective upregulation of mitochondrial functional gene clusters plays a more prominent role in the production process of 2-PE. Finally, findings from untargeted metabolomics suggested that by enhancing respiratory metabolism and reducing the Crabtree effect, the accumulation of metabolites resisting high 2-PE stress was observed, such as intracellular amino acids and purines. Hence, this strategy provided a richer supply of precursors and cofactors, effectively promoting the synthesis of 2-PE. In short, this study provides a bridge for studying the metabolic mechanism of high-yield 2-PE strains with the subsequent targeted strengthening of relevant synthetic pathways. It also provides insights for the synthesis of nonalcoholic products in S. cerevisiae.

Promoter screening and identification for metabolic regulation in Acremonium chrysogenum.

Acremonium chrysogenum is the major industrial producer of cephalosporin C (CPC), which is used as raw material for the production of significant cephalosporin antibiotics. Due to the lack of diverse promoter elements, the development of metabolic engineering transformation is relatively slow, resulting in a limited improvement on CPC production. In this study, based on the analysis of the transcriptome profile, 27 candidate promoters were selected to drive the expression of the reporter genes. The promoter activities of this library ranged from 0.0075 to 101 times of the control promoter PAngpdA . Simultaneously, a rapid screening method for potential bidirectional promoters was developed and 4 strong bidirectional promoters from 27 candidate options were identified and validated. Finally, the Golden Gate method was employed to combine promoter modules from the library with various target genes. Through a mixed transformation and screening process, high-yielding strains AG-6, AG-18, and AG-41 were identified, exhibiting an increase in CPC production of 30%, 35%, and 29%, respectively, compared to the control strain Ac-∆axl2:: eGFP. Therefore, the utilization of this promoter library offers a broader range of synthetic biology toolkits for the genetic engineering transformation of A. chrysogenum, thus establishing a solid foundation for the precise regulation of gene expression.

Evaluation of a Virus-like Nanoparticle Porcine Circovirus Type-2 (PCV2) Capsid Protein Fused with the Pig Immunoglobulin Fc Fragment as a Novel Vaccine Candidate against PCV2 in Mice.

Porcine circovirus Type 2 (PCV2) is a primary etiological pathogen of post-weaning multi-systemic wasting syndrome (PMWS). The capsid protein of PCV2 is the crucial immunogenic protein which can induce antibody generation and immune responses. However, there is still a lack of efficient PCV2 vaccines with high immunogenicity. In the current study, we developed a novel engineered PCV2 capsid (∆1-41aa)-pFc fusion protein (PCFP), which comprised a truncated capsid protein of PCV2 and a porcine IgG Fc fragment, fused to the capsid protein of PCV2 at the C-terminus. We found that this novel fusion protein could auto-assemble into virus-like nanoparticles with an estimated mean diameter of 22.6 nm, characterized by transmission electron microscopy. Immunization of BALB/c mice with this fusion protein significantly increased the production levels of anti-PCV2-capsid protein antibody in serum. Besides, the virus-like nanoparticles, PCFP was demonstrated to induce efficient cellular immune responses in mice, as evident by the high specific T cell reactivity to the PCFP fusion protein and the high production of the immune cytokines IFN-γ and IL-10 in an ex vivo re-stimulation system. Collectively, these findings demonstrate that the PCV2 truncated capsid subunit Fc-fusion protein can induce both cellular and humoral immune responses, and it displays great application potential.

Rationally optimized generation of integrated Escherichia coli with stable and high yield lycopene biosynthesis from heterologous mevalonate (MVA) and lycopene expression pathways.

The stability and high productivity of heterogeneous terpenoid production in Escherichia coli expression system is one of the most key issues for its large scale industrialization. In the current study on taking lycopene biosynthesis as an example, an integrated Escherichia coli system has been generated successfully, which resulted into stable and high lycopene production. In this process, two modules of mevalonate (MVA) pathway and one module of lycopene expression pathway were completely integrated in the chromosome. Firstly, the copy number and integrated position of three modules of heterologous pathways were rationally optimized. Later, a strain DH416 equipped with heterogeneous expression pathways through chromosomal integration was efficiently derived from parental strain DH411. The evolving DH416 strain efficiently produced the lycopene level of 1.22 g/L (49.9 mg/g DCW) in a 5 L fermenter with mean productivity of 61.0 mg/L/h. Additionally, the integrated strain showed more genetic stability than the plasmid systems after successive 21st passage.

Metabolic engineering coupled with adaptive evolution strategies for the efficient production of high-quality L-lactic acid by Lactobacillus paracasei.

The main indicators for industrial production of high-quality lactic acid at elevated temperatures are high titer, productivity, yield, and optical purity. However, no such strains have been reported to meet all these requirements simultaneously. In this study, a high optical purity L-lactic acid producing strain is developed through the CRISPR-Cas9 gene editing platform. Further, adaptive evolution was used to breed and select a high-performance strain (NCBIO01-M2-ldhL1-HT) that could efficiently produce L-lactic acid at a high temperature of 45℃. This strain produced 221.0 g/L of L-lactic acid in open fermentation with high initial glucose concentration. Also, L-lactic acid productivity and yield was above 7.5 g/L/h and 0.96 g/g respectively, as well as the optical purity of L-lactic acid in the fermentation broth exceeded 99.1%. In short, this breeding strain possess high potential to be considered for the commercial production of polymer-grade L-lactic acid.

Exploring the metabolic fate of propanol in industrial erythromycin-producing strain via 13C labeling experiments and enhancement of erythromycin production by rational metabolic engineering of Saccharopolyspora erythraea.

Propanol had been widely used as a precursor for erythromycin synthesis in industrial production. However, the knowledge on the exact metabolic fate of propanol was still unclear. In the present study, the metabolic fate of propanol in industrial erythromycin-producing strain Saccharopolyspora erythraea E3 was explored via 13C labeling experiments. An unexpected pathway in which propanol was channeled into tricarboxylic acid cycle was uncovered, resulting in uneconomic catabolism of propanol. By deleting the sucC gene, which encodes succinyl-CoA synthetase that catalyse a reaction in the unexpected propanol utilization pathway, a novel strain E3-ΔsucC was constructed. The strain E3-ΔsucC showed a significant enhancement in erythromycin production in the chemically defined medium compared to E3 (786.61 vs 392.94 mg/L). Isotopically nonstationary 13C metabolic flux analysis were employed to characterize the metabolic differences between Saccharopolyspora erythraea E3 and E3-ΔsucC. The results showed that compared with the starting strain E3, the fluxes of pentose phosphate pathway in E3-△sucC increased by almost 200%. The flux of the metabolic reaction catalyzed by succinyl-CoA synthetase in E3-ΔsucC was almost zero, while the glyoxylate bypass flux significantly increased. These new insights into the precursor utilization of antibiotic biosynthesis by rational metabolic engineering in Saccharopolyspora erythraea provided the new vision in increasing industrial production of secondary metabolites.

[Study on changes of coagulation state, microthrombus and vascular density in the model of iron accumulation osteoporosis].

OBJECTIVE: To understand changes of coagulation state, microthrombus, microvascular bed and bone density in the osteoporosis model of iron accumulation, and explore the influence of iron accumulation in aspects of osteoporosis on coagulation function and blood vessels. METHODS: Tewnty-four male SPF SD rats aged 6 months were selected, which with the average body weight (250±20) g, which were divided into control group and iron accumulation group according to random number table, 12 rats in each group. Iron accumulation group was intervened by intraperitoneal injection of ferric ammonium citrate 90 mg / kg, and control group was intraperitoneally injected with equal volume of normal saline, twice a week for 9 weeks. After intervention, serum ferritin, coagulation function, microthrombus, vascular density, and three-dimensional morphological reconstruction and spatial structure parameters of the distal femur trabeculae were measured and statistically analyzed. RESULTS: Serum ferritin of iron accumulation group (136.36±35.41) µg / L was higher than control group (68.44±16.86) µg / L(P<0.05). Bone mineral density (BMD) of iron accumulation group (0.167±0.024) g / cm3 was lower than control group(0.400±0.030)g / cm3. Fibrinogen of iron accumulation group (2.03±0.13) g / L was increased than that of control group (1.78±0.46) g / L, D-dimer contents of iron accumulation group (534.95±31.81) ng /ml was increased than that of control group (329.02±84.99) ng /ml, while thrombin time (39.64±2.18) s and prothrombin time(8.70±0.39) s of iron accumulation group were shorter than that of control group (44.92±2.98) s, (9.44±0.49) s (P<0.05). After ink staining, microvessel density in iron accumulation group (17.46±2.07)% was significantly reducedcompared with that of control group(23.81±2.98)%(P<0.05). HE and MSB staining which showed microthrombus in bone marrow of iron accumulation rats, as well as microthrombus in myocardium. CONCLUSION: In the osteoporosis model with the influence of iron accumulation, iron accumulation had a significant influence on the coagulation function, and the blood was relatively hypercoagulable. The bone vascular bed uas reduced, and there were microthrombus in the bone marrow. Hypercoagulable state of blood and formation of microthrombi may be important factors influencing the occurrence of iron accumulation osteoporosis.

Bone Microthrombus Promotes Bone Loss in Iron Accumulation Rats.

In the present study, we investigated the changes of the coagulation state, bone microthrombus, microvascular bed and bone density levels in iron accumulation rats. Meanwhile,the effect of anticoagulation therapy on bone mineral density was further investigated. We established two groups: a control (Ctrl) group and an iron intervention (FAC) group. Changes in coagulation function, peripheral blood cell counts, bone microthrombus, bone vessels and bone mineral density were compared between the two groups. We designed the non-treatment group and treatment group to study the changes of bone mineral density by preventing microthrombus formation with the anticoagulant fondaparinux. We found that the fibrinogen and D-dimer contents were significantly higher, whereas the thrombin time (TT) and prothrombin time (PT) were significantly shorter in the FAC group. After ink staining, the microvascular bed in the FAC group was significantly reduced compared with that in the Ctrl group. HE and Martius Scarlet Blue (MSB) staining showed microthrombus in the bone marrow of the iron accumulation rats. Following anticoagulation therapy, the bone microcirculation vascular bed areas in the treatment group rats were significantly increased. Furthermore, the bone mineral density was increased in the treatment group compared with that in the non-treatment group. Through experiments, we found that the blood in iron accumulation rat was relatively hypercoagulable; moreover, there was microthrombus in the bone marrow, and the bone vascular bed was reduced. Additionally, anticoagulation was helpful for improving bone microcirculation, reducing microthrombus and decreasing bone loss.

Target-site directed rational high-throughput screening system for high sophorolipids production by Candida bombicola.

In this study we have established a rational and high performance high-throughput screening system to select for a sophorolipids (SLs) high-producing strain of Candida bombicola. Introduction of mutagen combination, relaxation culture and multi-stress significantly improved both mutation and positive mutation rates. A high-performing strain, Ncbio 5, was selected out of 6212 mutants. Final SLs titer, productivity and yield of this strain in 5 L bioreactors were 26.9%, 27.0% and 35.0% higher than in the original strain, respectively. The improved fermentation performance in Ncbio 5 is contributed by the longer production period, higher SLs productivity and more efficient oil utilization. The strategy adopted herein to optimize the high-throughput screening system should be readily extendable to other similar systems.

Xanthan-Curdlan nexus for synthesizing edible food packaging films.

In this study, we prepared a series of edible blend films of xanthan and curdlan by mixing different ratios of these two biopolymers. Characterization techniques like FTIR, XRD, TGA and SEM analysis were applied to investigate the newly formed films. Moreover, mechanical properties, moisture absorbance properties and water solubility of these films were also determined. The obtained results demonstrated that the strong intermolecular hydrogen bonding was observed between xanthan and curdlan at pH 5. At this pH, the xanthan-curdlan hydrogel retained the original structure of xanthan and sustained self-aggregation of xanthan chains via hydrogen bonding, which led to strong intermolecular bonding between xanthan and curdlan. Furthermore, the 5:5 and 4:6 ratios of xanthan and curdlan showed greater interaction in the blend films that resulted in their excellent miscibility. Moreover, highest tensile strength of 28.13 and 26.45 MPa were also found in the same rational of XG/CG blend films. In addition, it was observed that the curdlan incorporation improved the water solubility properties of XG/CG blend films. Conclusively, this xanthan/curdlan nexus with excellent mechanical and moisture barrier properties confirm its potential application and prospective use as food packaging material.

Mapping molecular pathways for embryonic Sertoli cells derivation based on differentiation model of mouse embryonic stem cells.

BACKGROUND: Embryonic Sertoli cells (eSCs) have been known for playing important roles in male reproductive development system. In current studies, eSCs were mainly generated from induced intermediate mesoderm. The deriving mechanism of eSCs has been unclear so far. Therefore, this work was aimed to reveal the molecular pathways during derivation of eSCs. METHODS: In this scenario, a differentiation model from mouse embryonic stem cells (mESCs) to eSCs was established through spatiotemporal control of 5 key factors, Wilms tumor 1 homolog (Wt1), GATA binding protein 4 (Gata4), nuclear receptor subfamily 5, group A, member 1 (Nr5a1, i.e., Sf1), SRY (sex determining region Y)-box 9 (Sox9), doublesex, and mab-3 related transcription factor 1 (Dmrt1). To investigate the molecular mechanism, these key factors were respectively manipulated through a light-switchable (light-on) system, tetracycline-switchable (Tet-on) system, and CRISPR/Cas9 knock out (KO) system. RESULTS: Via the established approach, some embryonic Sertoli-like cells (eSLCs) were induced from mESCs and formed ring-like or tubular-like structures. The key factors were respectively manipulated and revealed their roles in the derivation of these eSLCs. Based on these results, some molecular pathways were mapped during the development of coelomic epithelial somatic cells to eSCs. CONCLUSIONS: This differentiation model provided a high controllability of some key factors and brought a novel insight into the deriving mechanism of Sertoli cells.

Inducing Non-genetically Modified Induced Embryonic Sertoli Cells Derived From Embryonic Stem Cells With Recombinant Protein Factors.

Embryonic Sertoli cells (eSCs) possess multiple supporting functions and research value in gonadal development and sex determination. However, the limitation of acquiring quality eSCs had hindered the further application. Herein, we successfully derived non-genetically modified (non-GM)-induced embryonic Sertoli-like cells (eSLCs) from mouse embryonic stem cells (ESCs) with a TM4 cell-derived conditioned medium containing recombinant endogenous protein factors Sry, Sox9, Sf1, Wt1, Gata4, and Dmrt1. These eSLCs were determined through morphology; transcriptional expression levels of stage-specific, epithelial, and mesenchymal marker genes; flow cytometry, immunofluorescence; and immunocytochemistry and functionally determined by coculture with spermatogonia stem cells. Results indicated that these eSLCs performed similarly to eSCs in specific biomarkers and expression of marker genes and supported the maturation of spermatogonia. The study induced eSLCs from mouse ESCs by defined protein factors. However, the inducing efficiency of the non-GM method was still lower than that of the lentiviral transduction method. Thus, this work established a foundation for future production of non-GM eSLCs for clinical applications and fundamental theory research.

Combined available nitrogen resources enhanced erythromycin production and preliminary exploration of metabolic flux analysis under nitrogen perturbations.

In the current study, the effect of different available nitrogen sources on erythromycin fermentation by Saccharopolyspora erythraea No. 8 is evaluated. Three different combinations of corn steep liquor and yeast powder were developed to investigate their impacts on erythromycin production. The results indicate that the optimal combination of available nitrogen sources was 10.0 g/L corn steep liquor and 4.0 g/L yeast power, generating a maximum yield of erythromycin of 13672 U/mL. To explore the effects of nitrogen perturbations on cell metabolism, metabolic flux analyses were performed and compared under different conditions. A high flux pentose phosphate pathway provided more NADPH for erythromycin synthesis via nitrogen optimization. Moreover, high n-propanol specific consumption rate enhanced erythromycin synthesis and n-propanol flowed into the central carbon metabolism by methylmalonyl-CoA node. These results indicate that the selection of an appropriate organic nitrogen source is essential for cell metabolism and erythromycin synthesis, and this is the first report of the successful application of available nitrogen source combinations in industrial erythromycin production.

Efficient generation of male germ-like cells derived during co-culturing of adipose-derived mesenchymal stem cells with Sertoli cells under retinoic acid and testosterone induction.

BACKGROUND: Adipose-derived mesenchymal stem cells (ADMSCs) are considered an efficient and important candidate for male infertility treatment because they contain pluripotent stem cells, which can differentiate into all cells from three germ layers. However, the efficient generation of male germ-like cell (MGLCs) is one of the key issues, and little is known about the mechanisms underlying generation of MGLCs. Herein, we attempt to improve the efficient generation of MGLCs derived during co-culturing of rat ADMSCs with SCs under retinoic acid (RA) and testosterone (T) treatment. METHODS: ADMSCs isolated from male SD rat were induced into generation of MGLCs by using respective methods in vitro. Transwell insert system was used for co-culturing. Busulfan-induced non-obstructive azoospermia rat mode was used to evaluate spermatogenic recovery ability of treated ADMSCs. Besides, the relative gene expression level was detected by reverse transcription PCR, quantitative RT-PCR. The relative protein expression level was detected by western blot (WB) and immunostaining analysis. RESULTS: The results showed that ADMSCs co-cultured with TM4 cells under RA and T induction enhanced the formation of bigger and tightly packed MGLCs feature colonies in vitro. Moreover, the expression of male germ cell-related markers (Oct4, Stella, Ddx4, Dazl, PGP9.5, Stra8, and ITGα6) is significantly upregulated in TM4 cell-co-cultured ADMSCs in vitro and in busulfan-treated rat testis after injecting TM4 cell-treated ADMSCs for 2 months. Comparatively, the ADMSCs treated by TM4 cell with RA and T exhibited the highest expression of male germ cell-related markers. RA- and T-treated TM4 cell showed fewer dead cells and higher cytokine secretion than untreated groups. The protein expression level of TGFß-SMAD2/3, JAK2-STAT3, and AKT pathways in ADMSCs co-cultured with TM4 cells under RA and T was higher than others. Whereas, downregulation of male germ cell-related marker expression subsequently inhibited the phosphorylation of SMAD2/3, JAK2, STAT3, and AKT. CONCLUSION: These results suggested that TM4 cells could efficiently stimulate in vitro generation of MGLCs during co-culturing of ADMSCs under RA and T treatment. Conclusively, the ADMSCs co-cultured with TM4 cell under RA and T induction stimulate the efficient generation of MGLCs in vitro through activating TGFß-SMAD2/3, JAK2-STAT3, and AKT pathways. Among them, JAK2-STAT3 and AKT pathways are being first reported to show involvement of in vitro generation of MGLCs during ADMSC co-culturing with SCs.

Differentiation roadmap of embryonic Sertoli cells derived from mouse embryonic stem cells.

BACKGROUND: Embryonic Sertoli cells (eSCs) play an important role in sex determination and in male gonad development which makes them a very useful cell type for therapeutic applications. However, the deriving mechanism of Sertoli cells has been unclear and challenging to create a large number of quality eSCs. Therefore, this study aimed to create the eSCs induced from mouse embryonic stem (mES) cells by regulating defined factors and to explore the relevant regulatory mechanism. METHODS: Six inducing factors, Sry, Sox9, SF1, WT1, GATA4, and Dmrt1, were respectively transduced into mES cells by lentiviral infection according to the experimental design. The test groups were identified by development stage-specific markers, AMH, Emx2, SF1, and FasL, using flow cytometry. Induced eSCs were determined by FasL and AMH biomarkers under immunofluorescence, immunocytochemistry, and flow cytometry. Moreover, the pluripotency markers, gonad development-related markers, epithelial markers and mesenchymal markers in test groups were transcriptionally determined by qPCR. RESULTS: In this study, the co-overexpression of all the six factors effectively produced a large population of eSCs from mES cells in 35 days of culturing. These eSCs were capable of forming tubular-like and ring-like structures with functional performance. The results of flow cytometry indicated that the upregulation of GATA4 and WT1 contributed to the growth of somatic cells in the coelomic epithelium regarded as the main progenitor cells of eSCs. Whereas, SF1 facilitated the development of eSC precursor cells, and Sry and Sox9 promoted the determination of male development. Moreover, the overexpression of Dmrt1 was essential for the maintenance of eSCs and some of their specific surface biomarkers such as FasL. The cellular morphology, biomarker identification, and transcriptomic analysis aided in exploring the regulatory mechanism of deriving eSCs from mES cells. CONCLUSION: Conclusively, we have elucidated a differentiation roadmap of eSCs derived from mES cells with a relevant regulatory mechanism. Through co-overexpression of all these six factors, a large population of eSCs was successfully induced occupying 24% of the whole cell population (1 × 105 cells/cm2). By adopting this approach, a mass of embryonic Sertoli cells can be generated for the purpose of co-culture technique, organ transplantation, gonadal developmental and sex determination researches.

Sustainable biosynthesis of curdlan from orange waste by using Alcaligenes faecalis: A systematically modeled approach.

This study presents an engineered approach for sustainable biosynthesis of curdlan by Alcaligenes faecalis using orange peels. To confirm the substrate suitability a four step study was organized. Firstly, drying of substrate was carried within temperature range of 60-120 °C, along with the application of moisture diffusion control model. Secondly, fermentation medium was obtained via saccharification and detoxification, releasing highest sugar at 72.34 g/L with phenolics removal of 95-98%. Thirdly, curdlan fermentation was conducted in detoxified orange peel hydrolysate followed by optimization of batch culture fermentation via kinetic modeling using Logistic and Luedeking-Piret equations. In 5 L bioreactor, highest specific growth rate (µm = 0.233/h), highest curdlan production (Pm = 23.24 g/L) and growth associated rate constant (α = 3.403) were achieved. Moreover, the total sugar consumption and conversion rates were 83.27% and 53.20%. Lastly, characterization techniques such as FTIR, NMR, XRD, TGA, HPGPC and EDS were applied to biosynthesized curdlan for qualitative validation.

Kinetic analysis of sodium gluconate production by Aspergillus niger with different inlet oxygen concentrations.

To further understand fermentation kinetics of sodium gluconate (SG) production by Aspergillus niger with different inlet oxygen concentrations, logistic model for cell growth and two-step models for SG production and glucose consumption were established. The results demonstrated that the maximum specific growth rate (µm) presented exponential relationship with inlet oxygen concentration and the maximum biomass (Xm) exhibited linear increase. In terms of SG production, two-step model with Luedeking-Piret equation during growth phase and oxygen-dependent equation during stationary phase could well fit the experimental data. Notably, high inlet oxygen concentration exponentially improved SG yield (YP/S), whereas biomass yield to glucose (YX/S) and cell maintenance coefficient (m) were almost independent on inlet oxygen concentration, indicating that high oxygen supply enhancing SG synthesis not only functioning as a substrate directly, but also regulating glucose metabolism towards SG formation. Finally, the applicability and predictability of the proposed models were further validated by additional experiments.

Co-culture with TM4 cells enhances the proliferation and migration of rat adipose-derived mesenchymal stem cells with high stemness.

The proliferation and migration of mesenchymal stem cells (MSCs) are the efficiency determinants in MSCs transplant therapy. Sertoli cells considered as "nurse cell" possesses the ability to enhance the proliferation and migration of umbilical cord mesenchymal stem cells (UCMSCs). However, no reports about TM4 cells' effect on the proliferation and migration of adipose tissue-derived mesenchymal stem cells (ADSCs) have been found until at present research work. Therefore, this study investigates the effect of TM4 cells on the proliferation and migration of ADSCs. We found that the performance of proliferation and migration of ADSCs were improved significantly while maintaining their stemness and reducing their apoptosis rate. After co-culturing with TM4 cells, the co-cultured ADSCs demonstrated higher proportion of synthetic phase (S) cells and colony-forming units-fibroblastic (CFU-F) number, lower proportion of sub-G1 phase cells and enhanced osteogenic and adipogenic differentiation ability. Moreover, results confirmed the higher multiple proteins involved in cell proliferation and migration including expression of the phospho-Akt, mdm2, pho-CDC2, cyclin D1 CXCR4, MMP-2, as well as phospho-p44 MAPK and phospho-p38 MAPK in co-cultured ADSCs. Furthermore, the process of TM4 cells promoting the proliferation of ADSCs was significantly inhibited by the administration of the PI3K/AKT inhibitor LY294002. Obtained results indicated that TM4 cells through MAPK/ERK1/2, MAPK/p-38 and PI3K/Akt pathways influence the proliferation and migration of ADSCs. These findings indicated that TM4 cells were found effective in promoting stemness and migration of ADSCs, that proves adopted co-culturing technique as an efficient approach to obtain ADSCs in transplantation therapy.

Kinetic analysis of curdlan production by Alcaligenes faecalis with maltose, sucrose, glucose and fructose as carbon sources.

Curdlan has wide-ranging benefits in food and pharmaceutical industries for its unique rheological and thermal gelling properties. To analyze the cell growth and curdlan biosynthesis kinetics of Alcaligenes faecalis, the kinetic properties of the curdlan fermentation under different carbon sources conditions (maltose, sucrose, glucose and fructose) were investigated using Logistic and Luedeking-Piret equations. The results demonstrated that curdlan fermentation is partial growth-associated process. With maltose as the sole carbon source, the highest curdlan production (Pm = 39.3 g/L), the maximum specific growth rate (µm = 0.44/h) and the growth-associated rate constant (α = 2.05 g curdlan/g cell) were achieved. In contrast, the fructose was the less desired carbon source in both the cell growth and curdlan production. Further, the results demonstrated that slow-releasing glucose from maltose boosted cell growth and curdlan production.