[Effect of topical glucocorticoid in treating phimosis on urinary tract infection of vesicoureteral reflux in infants].

Objective: To evaluate the effect of topical glucocorticoid in treating phimosis on urinarytract infection(UTI) of vesicoureteral reflux(VUR) in infants. Methods: Clinical data of infants with UTI diagnosed as primary VUR admitted to our hospital from January 2016 to January 2021 were retrospectively analyzed. The children were divided into three groups:the effective group (topical glucocorticoid was effective in the treatment of phimosis), the ineffective group(topical glucocorticoid was ineffective in the treatment of phimosis), and the untreated group(phimosis was not treated). Age of onset, degree of reflux, side and other indicators were compared to understand the effectiveness of topical glucocorticoid in treating phimosis, and the clinical characteristics of repeated UTI with VUR in treated phimosis and untreated phimosis. Results: A total of 544 children were included. Among them, 59 cases were treated with topical glucocorticoid for phimosis, 48 cases in the effective group, and their age was (12.5±8.4) months;11 cases in the ineffective group,and their age was (11.2±8.9) months. There were 485 cases in the untreated group, and their age was (13.1±9.3) months.The effective rate of topical glucocorticoid in the treatment of phimosis was 81.36%. There were 12 cases(12/48) of recurrent UTI in the effective group and 213 cases (213/485)of recurrent UTI in the untreated group, and the difference between the two groups was statistically significant (P=0.008). Conclusion: Treatment of phimosis with topical glucocorticoid is an effective, easy to perform, and cost-effective method, and can effectively reduce the risk of recurrent UTI in infants with primary VUR.

[Relationship between 25-hydroxyvitamin D levels and idiopathic benign paroxysmal positional vertigo in female patients].

Objective: To investigate the relationship between serum 25-hydroxyvitamin D[25(OH) D] levels and idiopathic benign paroxysmal positional vertigo (BPPV) in female patients. Methods: A total of 67 women diagnosed with idiopathic BPPV were selected as the study group between January and December 2016 in Ningbo No.2 Hospital, and 95 healthy women without a history of vertigo or dizziness were selected as the control group. The data of height, weight, histories of hypertension and diabetes mellitus were collected, and serum 25(OH) D levels were measured. The number of the recurrence and canalith repositioning maneuvers were recorded during the one-year follow-up. Results: No significant differences of age, body mass index, histories of hypertension and diabetes mellitus between patients with BPPV and healthy controls (all P>0.05) .The serum 25 (OH) D levels were significantly lower in patients with idiopathic BPPV than in healthy controls [(50.56±13.36) nmol/L vs (56.55±16.21) nmol/L, t=-2.485, P=0.014]. BPPV patients with low level of 25(OH) D showed a significant increase in the number of canalith repositioning maneuvers required and the recurrence rate. The regression analyses demonstrated that 25(OH)D deficiency was associated with BPPV with an odds ratio of 2.054 (95% CI: 1.088-3.877, P=0.026). Conclusion: 25(OH) D deficiency may be a risk factor of BPPV.

Efficacy and community effectiveness of larvivorous fish for dengue vector control.

OBJECTIVE: To evaluate the efficacy and community effectiveness of larvivorous fish for the control of dengue vectors and dengue transmission, when used as a single agent or in combination with other vector control methods. METHOD: Comprehensive literature search of published and grey literature using PubMed, EMBASE (DMDI), Web of Science, WHOLIS, WILEY, LILACS, GIFT, Cochrane Library, ELDIS, New York Academy of Medicine Grey Literature Report and Google. All results were checked for duplicates and examined for eligibility. Methodological quality of the studies was assessed using RoBANS. RESULTS: Thirteen articles were considered eligible for inclusion. Incorporating a wide range of interventions and outcome measures, three were efficacy studies and 10 assessed community effectiveness. None of the studies were randomised or cluster-randomised controlled trials. All three efficacy studies and seven community effectiveness studies investigated fish as a single agent. All efficacy studies reported elimination of Aedes larvae from treated containers, while community effectiveness studies reported reductions in immature vector stages, two of which also detected a continuous decline over 2 years. An impact on adult mosquitoes was shown in only two community effectiveness studies. Reductions in dengue cases following intervention were reported in two studies, but it was not possible to attribute this to the intervention. CONCLUSION: While the use of larvivorous fish as a single agent or in combination with other control measures could lead to reductions in immature vector stages, considerable limitations in all the studies restricted any conclusions with respect to the evaluation of community effectiveness. Evidence for the community effectiveness of larvivorous fish as a single agent remains minimal and cluster-randomised controlled studies that include the assessment of impact on dengue are recommended.

Community effectiveness of copepods for dengue vector control: systematic review.

OBJECTIVE: Vector control remains the only available method for primary prevention of dengue. Several interventions exist for dengue vector control, with limited evidence of their efficacy and community effectiveness. This systematic review compiles and analyses the existing global evidence for community effectiveness of copepods for dengue vector control. METHODS: The systematic review follows the PRISMA statement, searching six relevant databases. Applying all inclusion and exclusion criteria, 11 articles were included. RESULTS: There is evidence that cyclopoid copepods (Mesocyclops spp.) could potentially be an effective vector control option, as shown in five community effectiveness studies in Vietnam. This includes long-term effectiveness for larval and adult control of Ae. aegypti, as well as dengue incidence. However, this success has so far not been replicated elsewhere (six studies, three community effectiveness studies--Costa Rica, Mexico and USA, and three studies analysing both efficacy and community effectiveness--Honduras, Laos and USA), probably due to community participation, environmental and/or biological factors. Judging by the quality of existing studies, there is a lack of good study design, data quality and appropriate statistics. CONCLUSION: There is limited evidence for the use of cyclopoid copepods as a single intervention. There are very few studies, and more are needed in other communities and environments. Clear best practice guidelines for the methodology of entomological studies should be developed.

Long non-coding RNA ANCR promotes progression of NSCLC by inhibiting E-Ca expression.

The article "Long non-coding RNA ANCR promotes progression of NSCLC by inhibiting E-Ca expression, by T. Zhou, J.-J. Fang, Y.-X. Zhou, Z.-P. Li, L. Jiang, W.-W. Han, Z.-H. Zhu, published in Eur Rev Med Pharmacol Sci 2020; 24 (3): 1250-1257-DOI: 10.26355/eurrev_202002_20178-PMID: 32096155" has been withdrawn from the authors. They stated that "during the last few months, new experimental data have been obtained and analyzed, so we want to rewrite the paper to further test the cell experiment and give more evidence to support our paper. It is difficult to repeat the part of the cell experiment, and the results need to be further improved. In addition, the internal reference pictures in the paper are wrong and need to be re experimented and modified. This needs more detailed studies and maybe a long time, so we want to withdraw our paper and resubmit it when it is ready". The Publisher apologizes for any inconvenience this may cause. https://www.europeanreview.org/article/20178.

Long non-coding RNA ANCR promotes progression of NSCLC by inhibiting E-Ca expression.

OBJECTIVE: This study aimed to investigate whether long-chain non-coding ANCR is involved in the progression of non-small cell LCa (NSCLC) and its possible molecular mechanisms. PATIENTS AND METHODS: Quantitative Real Time-Polymerase Chain Reaction (qRT-PCR) was applied to examine ANCR expression in 48 cases of NSCLC and adjacent normal tissues. In addition, ANCR level in patients of different tumor staging was analyzed. The Kaplan-Meier method was applied to analyze the interplay between ANCR expression and the prognosis of patients with NSCLC. Subsequently, qRT-PCR was performed to detect ANCR level in LCa cell lines. After knocking down ANCR in A549 cells, ANCR and E-Ca mRNA expression were examined by qRT-PCR, while the expression levels of epithelial-mesenchymal transition (EMT)-related proteins were detected by Western blot. At the same time, cell viability and migration ability were analyzed through cell counting kit-8 (CCK-8) and cell wound healing assay, respectively. RNA immunoprecipitation (RIP) test was performed to verify the binding of ANCR to EZH2. After knocking down EZH2 in A549 cells, E-Ca messenger ribonucleic acid (mRNA) expression was detected. Additionally, Chromatin immunoprecipitation (ChIP) assay was performed to detect the binding of EZH2 to the E-Ca promoter region. When E-Ca and ANCR were simultaneously knocked down in A549 cells, Western blot investigation was performed to examine the expression of EMT-related proteins, while CCK-8 and wound healing assays were applied to figure out the changes in cell viability and cell migration capacity. RESULTS: ANCR level was conspicuously higher in NSCLC tissues than that in normal tissues, and that in T3 and T4 tumors was also higher than that in T1 and T2. Meanwhile, ANCR expression in the tissues of patients with lymph node metastasis was conspicuously higher than those without metastasis. Survival analysis revealed that the overall survival of patients with NSCLC with high expression of ANCR was conspicuously lower than patients with low expression of ANCR. The qRT-PCR study verified that ANCR was highly expressed in the LCa cell line A549. After knocking down ANCR in A549 cells, ANCR and E-Ca mRNA levels were found conspicuously decreased, and so were the expression levels of EMT-related proteins, as well as the cell viability and migration ability. The RIP assay result indicated that ANCR can indeed bind to EZH2. E-Ca mRNA expression was elevated after the knockdown of EZH2 in A549 cells. In addition, the result of CHIP test demonstrated that EZH2 could combine with E-Ca. Simultaneous down-regulation of ANCR and E-Ca in A549 cells could reverse the influence of knocking down ANCR alone on cell viability and migration ability. CONCLUSIONS: Long-chain non-coding RNA ANCR was highly expressed in NSCLC tissues and could enhance the viability and malignancy of NSCLC cells by inhibiting the expression of E-Ca, thereby promoting the progression of NSCLC.

Effect of miR-10a on sepsis-induced liver injury in rats through TGF-ß1/Smad signaling pathway.

OBJECTIVE: To observe the effect of the micro ribonucleic acid (miR)-10a on sepsis-induced liver injury in rats through the transforming growth factor-b1 (TGF-b1)/Smad signaling pathway. MATERIALS AND METHODS: The rat model of sepsis was established via cecal ligation and puncture, in which miR-10a was overexpressed and silenced using liposome transfection. The rats were randomly divided into miR-10a mimics group (Mimics group, n=10) and miR-10a inhibitors group (Inhibitors group, n=10), and the sham operation group (Sham group, n=10) was also set up. The transfection efficiency of miR-10a in liver tissues in each group was detected via quantitative Real Time-Polymerase Chain Reaction (qRT-PCR), the serum liver function indexes aspartate aminotransferase (AST), alanine aminotransferase (ALT) and lactate dehydrogenase (LDH) were determined. Moreover, the content of the serum reactive oxygen species (ROS), glutathione (GSH), and GSH peroxidase (GSH-Px) was determined using enzyme-linked immunosorbent assay (ELISA). The content of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and myeloperoxidase (MPO) in liver tissues was detected, and the pathological changes in liver tissues were observed through hematoxylin-eosin (HE) staining. Finally, the expression levels of cytochrome P4502E1 (CYP2E1) and TGF-b1/Smad signaling pathway genes and proteins in liver tissues were detected via qRT-PCR and Western blotting. RESULTS: The expression of miR-10a was significantly increased in Mimics group (p<0.05) and extremely low in the Inhibitors group (p<0.05). In Mimics group, the levels of serum AST, ALT, and LDH were significantly increased (p<0.05), the content of ROS, TNF-α, IL-6, and MPO was substantially increased (p<0.05), while that of GSH and GSH-Px notably declined (p<0.05). According to the HE staining results, the liver cells were orderly arranged in the Inhibitors group, and they were disorderly arranged with more inflammatory cells in the Mimics group. The results of the gene and protein assays showed that the expression levels of CYP2E1, TGF-b1, and Smad2 in Mimics group were markedly higher than those in the Sham group (p<0.05), while they displayed the opposite trends in the Inhibitors group (p<0.05). CONCLUSIONS: Silencing miR-10a can inhibit the occurrence of sepsis-induced liver injury in rats by downregulating the TGF-ß1/Smad pathway.

[Study on bone metabolism in postmenopausal women with idiopathic benign paroxysmal positional vertigo].

Objective: To analyze the bone mineral density and serum osteocalcin levels in postmenopausal women with idiopathic benign paroxysmal positional vertigo. Methods: A total of 64 postmenopausal women with idiopathic BPPV were selected as the study group, and 98 postmenopausal healthy women were selected as the control group. Bone mineral density and serum osteocalcin levels were analyzed and compared between the groups.χ(2) test was used for numeration data and t test was used for measurement data. Results: The prevalence of osteoporosis or osteopenia in BPPV group 67.2% (43/64) was significantly higher than that in the control group 51.0% (50/98) (χ(2)=4.139, P=0.042). Among BPPV subjects, there was a negative correlation between osteocalcin and bone density T (r=-0.469, P<0.001). Osteocalcin was found in normal bone mass subgroup (13.61±4.32)µg/L, decreased bone mass subgroup (17.49±7.61)µg/L, and osteoporosis subgroup (20.83±6.72)µg/L, respectively, and the difference was statistically significant (F=5.39, P=0.007). Conclusions: Bone mineral density in BPPV group is lower than that in control group. The lower the bone mineral density of the patients, the higher the osteocalcin in BPPV group.

Kinetics and mechanisms of hydrolysis of 1,4-benzodiazepines I: chlordiazepoxide and demoxepam.

Differential absorbance spectroscopy was successfully used to follow the hydrolysis kinetics of chlordiazepoxide and demoxepam from pH 1 to 11. Loss of the methylamino group from chlordiazepoxide produced demoxepam. Demoxepam degraded by a parallel consecutive reaction to 2-amino-5-chlorobenzophenone and a glycine derivative. Two intermediates were observed by TLC for demoxepam hydrolysis. One was assigned the open-ring structure resulting from amide hydrolysis, which kinetically appears to be the major mechanistic route leading to the benzophenone product. The other intermediate, representing an alternative but minor pathway, presumably results from initial scission of the azomethine linkage. Protonation of the N-oxide slightly alters the importance of these two pathways. Recyclization of the carboxylic acid intermediate was facile at pH values below the pKa of this intermediate. The stability parameters involving buffer catalysis, ionic strength effects, and temperature dependence of rate constants are reported.

Kinetics and mechanisms of hydrolysis of 1,4-benzodiazepines II: oxazepam and diazepam.

The hydrolysis kinetics of oxazepam and diazepam leading to a benzophenone product and a glycine derivative were quantified from pH 1 to 11. For oxazepam, two intermediates were isolated and identified, indicating a parallel consecutive reaction mechanism. The hydrolysis occurred uncatalyzed and demonstrated acid-base catalysis for both reaction steps. One intermediate was observed by TLC for diazepam hydrolysis. This intermediate, resulting from breakage of the azomethine linkage, was different than the major intermediate isolated for oxazepam hydrolytic degradation (amide hydrolysis preferred). Stability parameters involving rate constant-temperature dependence are reported.

Kinetics and mechanisms of hydrolysis of 1,4-benzodiazepines III: nitrazepam.

The hydrolysis of nitrazepam involves a two-step sequential mechanism. The intermediate is the ring-opened compound resulting from scission of the azomethine bond. The final products are glycine and 2-amino-5-ni-robenzophenone. Recyclization of the intermediate to nitrazepam occurs at pH values above the pKa of the intermediate, in the pH region where the amino group of the intermediate is not protonated. As opposed to chlordiazepoxide and oxazepam, the initial hydrolysis step occurs at the 4,5-bond, not at the 1,2-amide linkage. This difference is attributed to a preferential activation for hydrolysis of the azomethine linkage by the nitro group. The hydrolysis involves an uncatalyzed reaction, specific acid-base catalysis, and general acid-base catalysis for acetate and phosphate buffers.

Revision stapedectomy: intraoperative findings, results, and review of the literature.

Seventy-four revision stapedectomies performed consecutively over 10 years (1986 to 1995) were reviewed retrospectively. The most common intraoperative findings were incus erosion, prosthesis displacement, and oval window closure. Incus erosion was more frequently associated with multiple revisions. The postoperative results were reported using the conventional method (postoperative air minus preoperative bone) as well as the guidelines recently published by the American Academy of Otolaryngology--Head and Neck Surgery (postoperative air minus postoperative bone), with success rates of postoperative air-bone gap closure to within 10 dB after revision surgery of 51.6% and 45.6%, respectively. Patients with persistent conductive hearing loss (large residual air-bone gaps) after primary stapedectomy had poorer postrevision hearing results. Sensorineural hearing loss (defined as a drop in bone pure-tone average of more than 10 dB) occurred in four cases (5.4%). The number of revision surgeries, variations in operative techniques using laser or drill, and the ossicle to which the prosthesis was attached did not statistically affect the postoperative air-bone gaps. These results were compared with previously published data.

[Studies on chemical structure of polysaccharide from fruit body of Coriolus versicolor].

AIM: To study the structure of polysaccharides from Coriolus versicolor. METHODS: By chromatography a homogeneous polysaccharide B-1-3 was obtained. The structure of B-1-3 was elucidated using the method of total acid hydrolysis, methylation, and 1D and 2D NMR. RESULTS: B-1-3 was found to be mainly composed of Glc by total acid hydrolysis. Its molecular weight was estimated to be 3.16 x 10(5) by HPLC method. Methylation analysis and 1H and 13CNMR indicated the main chain of B-1-3 consisted of beta-D-1,4-Glc and beta-D-1,3-Glc, and branch chains were situated, beta-D-1,3,6-Glc and beta-D-1,4,6-Glc. 1H and 13CNMR also showed that beta-D-Glc-1-->3-beta-D-Glc-beta-D-Glc-1-->4-beta-D-Glc and beta-D-Glc-1-->4-beta-D-Glc-beta-D-Glc-1-->4-beta-D-Glc fragments existed in B-1-3. CONCLUSION: B-1-3 is a glucan which main chain was composed of beta-D-1,4-Glc and beta-D-1,3-Glc beta-D-1,6-Glc.

Biological significance of nanograined/ultrafine-grained structures: Interaction with fibroblasts.

Given the need to develop high strength/weight ratio bioimplants with enhanced cellular response, we describe here a study focused on the processing-structure-functional property relationship in austenitic stainless steel that was processed using an ingenious phase reversion approach to obtain an nanograined/ultrafine-grained (NG/UFG) structure. The cellular activity between fibroblast and NG/UFG substrate is compared with the coarse-grained (CG) substrate. A comparative investigation of NG/UFG and CG structures illustrated that cell attachment, proliferation, viability, morphology and spread are favorably modulated and significantly different from the conventional CG structure. These observations were further confirmed by expression levels of vinculin and associated actin cytoskeleton. Immunofluorescence studies demonstrated increased vinculin concentrations associated with actin stress fibers in the outer regions of the cells and cellular extensions on NG/UFG substrate. These observations suggest enhanced cell-substrate interaction and activity. The cellular attachment response on NG/UFG substrate is attributed to grain size and hydrophilicity and is related to more open lattice in the positions of high-angle grain boundaries.

Cellular activity of bioactive nanograined/ultrafine-grained materials.

Our recent electron microscopy study on biomimetic nanostructured coatings on nanograined/ultrafine-grained (NG/UFG) substrates [Mater Sci Eng C 2009;29:2417-27] indicated that electrocrystallized nanohydroxyapatite (nHA) on phase-reversion-induced NG/UFG substrates exhibited a vein-type interconnected and fibrillar structure that closely mimicked the hierarchical structure of bone. The fibrillar structure on NG/UFG substrate is expected to be more favorable for cellular response than a planar surface. In contrast, hydroxyapatite (HA) coating on coarse-grained (CG) substrate more closely resembled a film rather than a fibrillar structure. Inspired by the differences in the structure of HA coating, we describe here the cell-substrate interactions of pre-osteoblasts (MC 3T3-E1) on bioactive NG/UFG and CG austenitic stainless steel substrates. NG/UFG austenitic stainless steel was obtained by a novel controlled phase-reversion annealing of cold-deformed austenite. This example provides an illustration of how a combination of cellular and molecular biology, materials science and engineering can advance our understanding of cell-substrate interactions. Interestingly, the cellular response of nanohydroxyapatite (nHA)-coated NG/UFG substrate demonstrated superior cytocompatibility, improved initial cell attachment, higher viability and proliferation, and well-spread morphology in relation to HA-coated CG substrate and their respective uncoated (bare) counterparts as implied by fluorescence and electron microscopy and MTT assay. Similar conclusions were derived from an immunofluorescence study that involved examination of the expression levels of vinculin focal adhesion contacts associated with dense actin stress fibers and fibronectin, protein analysis through protein bands in SDS-PAGE, and quantitative total protein assay. The enhancement of cellular response followed the sequence: nHA-coated NG/UFG>nHA-coated CG>NG/UFG>CG substrates. The outcomes of the study are expected to counter the challenges associated with the engineering of nanostructured surfaces with specific physical and surface properties for medical devices with significantly improved cellular response.

Biomimetic chitosan-nanohydroxyapatite composite scaffolds for bone tissue engineering.

We describe a comparative assessment of the structure-property-process relationship of three-dimensional chitosan-nanohydroxyapatite (nHA) and pure chitosan scaffolds in conjunction with their respective biological response with the aim of advancing our insight into aspects that concern bone tissue engineering. High- and medium-molecular-weight (MW) chitosan scaffolds with 0.5, 1 and 2 wt.% fraction of nHA were fabricated by freezing and lyophilization. The nanocomposites were characterized by a highly porous structure and the pore size (approximately 50 to 120 microm) was in a similar range for the scaffolds with different content of nHA. A combination of X-ray diffraction, Fourier transform infrared spectroscopy and electron microscopy indicated that nHA particles were uniformly dispersed in chitosan matrix and there was a chemical interaction between chitosan and nHA. The compression modulus of hydrated chitosan scaffolds was increased on the addition of 1 wt.% nHA from 6.0 to 9.2 kPa in high-MW scaffold. The water uptake ability of composites decreased with an increase in the amount of nHA, while the water retention ability was similar to pure chitosan scaffold. After 28 days in physiological condition, nanocomposites indicated about 10% lower degree of degradation in comparison to chitosan scaffold. The biological response of pre-osteoblasts (MC 3T3-E1) on nanocomposite scaffolds was superior in terms of improved cell attachment, higher proliferation, and well-spread morphology in relation to chitosan scaffold. In composite scaffolds, cell proliferation was about 1.5 times greater than pure chitosan after 7 days of culture and beyond, as implied by qualitative analysis via fluorescence microscopy and quantitative study through MTT assay. The observations related to well-developed structure morphology, physicochemical properties and superior cytocompatibility suggest that chitosan-nHA porous scaffolds are potential candidate materials for bone regeneration although it is necessary to further enhance the mechanical properties of the nanocomposite.

Clinical manifestations and outcomes of antithrombotic treatment of the Tan Tock Seng Hospital Singapore antiphospholipid syndrome cohort.

To examine the clinical manifestations, intensity of oral anticoagulation and outcomes in the prevention of recurrent thromboses in patients with antiphospholipid syndrome (APS) in a tertiary rheumatology centre in Singapore. Retrospective case review of consecutive patients with APS attending a rheumatology clinic from 1st January 2004 to 31st December 2005. There were 59 (44%) patients with definite APS and 75 (56%) with probable APS. Systemic lupus erythematosus (SLE) was the most common cause of secondary APS. Hypertension and hyperlipidaemia were the most common cardiovascular comorbidities. The most common manifestations were haematological (thrombocytopaenia and haemolytic anaemia), neurological (seizure, headache) and pulmonary hypertension. Among those with definite APS, there were similar proportions with arterial and venous thromboses. Recurrent thromboses occurred in 14 (23.7%) patient with definite APS receiving warfarin, comprising 14 (73.7%) episodes of arterial and 5 (26.3%) episodes of venous thromboses. Recurrent arterial thromboses occurred at international normalized ratio (INR) of <2 in 5 (35.7%), INR 2-3 in 6 (42.9%), INR > 3 in 3 (21.4%) episodes, respectively. Recurrent venous thromboses occurred at INR < 2 in 4 (80.0%) and INR > 3 in 1 (20.0%) episode, respectively. Twenty-eight episodes of bleeding occurred in 21 (35.6%) patients, the majority (78.6%) being minor bleeding. Two-thirds of all major bleeds occurred at INR >/= 3. Venous and arterial thromboses were equally common in our patients with definite APS, although recurrent thromboses were more common in the arterial circulation. Target INR > 3 was associated with lower rates of recurrent arterial thromboses but higher rates of major and recurrent bleeding. Target INR >/= 2 appeared to be sufficient to prevent recurrent venous thromboses.

Superior in vitro biological response and mechanical properties of an implantable nanostructured biomaterial: Nanohydroxyapatite-silicone rubber composite.

A potential approach to achieving the objective of favorably modulating the biological response of implantable biopolymers combined with good mechanical properties is to consider compounding the biopolymer with a bioactive nanocrystalline ceramic biomimetic material with high surface area. The processing of silicone rubber (SR)-nanohydroxyapatite (nHA) composite involved uniform dispersion of nHA via shear mixing and ultrasonication, followed by compounding at sub-ambient temperature, and high-pressure solidification when the final curing reaction occurs. The high-pressure solidification approach enabled the elastomer to retain the high elongation of SR even in the presence of the reinforcement material, nHA. The biological response of the nanostructured composite in terms of initial cell attachment, cell viability and proliferation was consistently greater on SR-5wt.% nHA composite surface compared to pure SR. Furthermore, in the nanocomposite, cell spreading, morphology and density were distinctly different from that of pure SR. Pre-osteoblasts grown on SR-nHA were well spread, flat, large in size with a rough cell surface, and appeared as a group. In contrast, these features were less pronounced in SR (e.g. smooth cell surface, not well spread). Interestingly, an immunofluorescence study illustrated distinct fibronectin expression level, and stronger vinculin focal adhesion contacts associated with abundant actin stress fibers in pre-osteoblasts grown on the nanocomposite compared to SR, implying enhanced cell-substrate interaction. This finding was consistent with the total protein content and SDS-PAGE analysis. The study leads us to believe that further increase in nHA content in the SR matrix beyond 5wt.% will encourage even greater cellular response. The integration of cellular and molecular biology with materials science and engineering described herein provides a direction for the development of a new generation of nanostructured materials.

Chitosan-gelatin scaffolds for tissue engineering: physico-chemical properties and biological response of buffalo embryonic stem cells and transfectant of GFP-buffalo embryonic stem cells.

The favorable cellular response of newly developed cell line, buffalo embryonic stem (ES) cells to three-dimensional biodegradable chitosan-gelatin composite scaffolds with regard to stem-cell-based tissue engineering is described. Chitosan-gelatin composites were characterized by a highly porous structure with interconnected pores, and the mechanical properties were significantly enhanced. Furthermore, X-ray diffraction study indicated increased amorphous content in the scaffold on the addition of gelatin to chitosan. To develop a transfectant of green fluorescence protein (GFP)-buffalo ES cell, transfection of GFP plasmid to the cell was carried out via the electroporation procedure. In comparison with pure chitosan, cell spreading and proliferation were greater in highly visualized GFP-expressing cell-chitosan-gelatin scaffold constructs. The relative comparison of biological response involving cell proliferation and viability on the scaffolds suggests that blending of gelatin in chitosan improved cellular efficiency. Studies involving scanning electron and fluorescence microscopy, histological observations and flow cytometer analysis of the constructs implied that the polygonal cells attached to and penetrated the pores, and proliferated well, while maintaining their pluripotency during the culture period for 28days. Chitosan-gelatin scaffolds were cytocompatible with respect to buffalo ES cells. The study underscores for the first time that chitosan-gelatin scaffolds are promising candidates for ES-cell-based tissue engineering.