The luteinizing hormone receptor (LHR) is a member of a subfamily of G protein-coupled receptors that is characterized by its alternative splicing. In a previous study, we identified a splice site mutation of intron 6 (IVS6-3C>A) in a patient suffering from Leydig cell hypoplasia, which leads to aberrant splicing of LHR mRNA. In vitro expression analysis confirmed that this mutation results in the skipping of exon 7 in the mature mRNA of the LHR gene. In this study, we determined the impact of IVS6-3C>A on the RNA secondary structure and function of LHR-Del7. The three-dimensional structure of the leucine-rich repeats in LHR was predicted by molecular modeling. Radioactive ligand-binding assays verified that LHR-Del7 has no binding affinity for hCG. Furthermore, we detected negligible cAMP production in cells transfected with LHR-Del7. Cells co-expressing LHR-WT and LHR-Del7 were able to generate cAMP in response to hCG, but there was no significant difference between cells transfected with LHR-WT/vector and LHR-WT/LHR-Del7, although the variant was able to localize to cell surface, similar to wild-type receptor. These results indicated that LHR-Del7 does not have a dominant negative effect on LHR-WT cell surface expression, and although the pathological splicing variant LHR-Del7 was able to localize to cell membranes it failed to bind hCG and had no effect on wild-type LHR.
function study; splicing variant; LHR
The title compound, C10H13ClN2O, was obtained as a by-product in the reaction of 2-chloromethyl-1H-benzimidazole, dimethyl sulfate and toluene to synthesise 2-chloromethyl-1-methylbenzimidazole. The dihedral angle between the benzene ring and the acetamide group is 89.72 (6)° while that between the aromatic ring and the chloracetyl group is 84.40 (4)°. In the crystal, adjacent molecules are linked by pairs of N—H⋯O hydrogen bonds into inversion dimers.
Multilocus sequence typing (MLST) based on six loci was used to analyze the relationship of 58 Candida tropicalis isolates from individual patients in a general hospital in Beijing, China. A total of 52 diploid sequence types (DSTs) were generated by the MLST, all of which were new to the central database. Unweighted Pair Group Method with Arithmetic Mean (UPGMA) dendrograms were constructed, which showed that the 58 isolates were distributed robustly and 6 main groups were clustered regardless of the specimen source and medical department. The minimum spanning tree (MST) of the 58 isolates (52 DSTs) and all 401 isolates (268 DSTs) in the C. tropicalis central database (http://pubmlst.org/ctropicalis/) indicated that the isolates in this study clustered in three relative pure clonal complexes, and 2 clustered with isolates from Taiwan, Belgium, Brazil, and the US. This study presents the first MLST analysis of C. tropicalis isolates from Mainland China, which may be useful for further studies on the similarity, genetic relationship, and molecular epidemiology of C. tropicalis strains worldwide.
During the past several years, multipotent mesenchymal stromal cells (MSCs) have rapidly moved from in vitro and animal studies into clinical trials as a therapeutic modality potentially applicable to a wide range of disorders. It has been proposed that ex vivo culture-expanded MSCs exert their tissue regeneration potential through their immunomodulatory and anti-inflammatory properties, and paracrine effects more than their ability to differentiate into multiple tissue lineages. Since extracellular matrix (ECM) deposition and tissue support is also one of many physiological roles of MSCs, there is increasing interest in their potential use for tissue engineering, particularly in combination with ECM-based scaffolds such as hyaluronic acid (HA). We investigated the effect of MSCs on immunophenotype of macrophages in the presence of an HA–hydrogel scaffold using a unique 3D coculture system. MSCs were encapsulated in the hydrogel and peripheral blood CD14+ monocyte-derived macrophages plated in direct contact with the MSC-gel construct. To determine the immunophenotype of macrophages, we looked at the expression of cell surface markers CD14, CD16, CD206, and human leukocyte antigen (HLA)-DR by flow cytometry. MSCs and macrophages cultured on the HA-hydrogel remained viable and were able to be recovered from the construct. There was a significant difference in the immunophenotype observed between monocyte-derived macrophages cultured on the HA scaffold compared to tissue culture polystyrene. Macrophages cultured on gels with MSCs expressed lower CD16 and HLA-DR with higher expression of CD206, indicating the least inflammatory profile overall, compatible with the immunophenotype of alternatively activated macrophages. Development of macrophages, with this immunophenotype, upon interaction with the MSC-hydrogel constructs may play a potentially significant role in tissue repair when using a cellular-biomaterial therapeutic approach.
When purified from a tumor, certain heat shock protein 70 (HSP70)-peptide complexes (PCs) can function as effective vaccines against the tumor from which the complexes were isolated. The immunogenic mechanisms of HSP70 preparations imply that tumor-derived HSP70-PCs exhibit antigens associated with antigen-presenting cells such as dendritic cells (DCs), inducing antigen-specific cytotoxic CD8+ T cells. However, some important membrane-resident tumor-associated peptides, such as the HER-2/neu (c-erbB2) oncogenic protein, cannot be purified from HSP70 by traditional methods. In the present study, a new approach for the purification of HSP70-PCs from HER-2-overexpressing breast cancer cells was established. The detergent 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS) was used to obtain more effectual tumor peptides. The new purified product was named HSP70-HER-2-PC, and its immunological activities were determined. Traditionally purified HSP70-PCs (without CHAPS) and recombinant human HSP70-HER-2 protein complexes (recombined in vitro) were used as controls. These three HSP70-associated tumor antigenic complex pulsed dendritic cells (DCs) were used to stimulate an antitumor response. The mature DCs pulsed with HSP70-HER-2-PCs stimulated autologous T cells to secrete higher levels of type I cytokine compared to the two control groups. Moreover, DCs pulsed with HSP70-HER-2-PCs induced the most specific CD8+ T cells that specifically killed the same tumor cells. These findings provide a basis for new approaches in enhancing HSP70-based immunotherapy for HER-2-associated or other membrane antigenic peptide-related cancers.
heat shock protein 70-peptide complexes; HER-2/neu oncogene protein; membrane protein; dendritic cells; CD8+ T cells; immunotherapy; cancer
Genetic polymorphisms of the Optic atrophy 1 gene have been implicated in altering the risk of primary open angle glaucoma (POAG), especially the susceptibility to normal tension glaucoma (NTG), but the results remain controversial.
Multiple electronic databases (up to January 20, 2012) were searched independently by two investigators. A meta-analysis was performed on the association between Optic atrophy 1 polymorphisms (rs 166850 and rs 10451941) and normal tension glaucoma (NTG)/high tension glaucoma (HTG). Summary odds ratios (ORs) and 95% confidence intervals (CI) were estimated.
Seven studies of 713 cases and 964 controls for NTG and five studies of 1200 cases and 971 controls for HTG on IVS8+4C>T (rs 166850) and IVS8+32T>C (rs10451941) were identified. There were significant associations between the OPA1 rs10451941polymorphism and NTG susceptibility for all genetic models(C vs. T OR = 1.26, 95% CI 1.09–1.47, p = 0.002; CC vs. TT: OR = 1.52, 95% CI 1.04–2.20, p = 0.029; CC vs. CT+TT: OR = 1.64, 95% CI 1.16–2.33, p = 0.005; CC+CT vs. TT: OR = 1.21, 95% CI 1.02–1.44, p = 0.032). However, no evidence of associations was detected between the OPA1 IVS8+32C>T polymorphism and POAG susceptibility to HTG. Similarly, clear associations between the rs 166850 variant and NTG were observed in allelic and dominant models (T vs. C OR = 1.52, 95% CI 1.16–1.99, p = 0.002; TT+TC vs. CC OR = 1.50, 95% CI 1.13–2.01, p = 0.006) but not to HTG. In subgroup analyses by ethnicity, we detected an association between both OPA1 polymorphisms and risk for NTG in Caucasians but not in Asians. By contrast, no significant findings were noted between OPA1 variants for HTG, either in Caucasians or in Asians.
Both the IVS8+4C>T and IVS8+32T>C variants may affect individual susceptibility to NTG. Moreover, stratified analyses for NTG detecting the effects of both OPA1 polymorphisms seemed to vary with ethnicity. Further investigations are needed to validate the association.
In China, rubella vaccination was introduced into the national immunization program in 2008, and a rubella epidemic occurred in the same year. In order to know whether changes in the genotypic distribution of rubella viruses have occurred in the postvaccination era, we investigate in detail the epidemiological profile of rubella in China and estimate the evolutionary rate, molecular clock phylogeny, and demographic history of the predominant rubella virus genotypes circulating in China using Bayesian Markov chain Monte Carlo phylodynamic analyses. 1E was found to be the predominant rubella virus genotype since its initial isolation in China in 2001, and no genotypic shift has occurred since then. The results suggest that the global 1E genotype may have diverged in 1995 and that it has evolved at a mutation rate of 1.65 × 10−3 per site per year. The Chinese 1E rubella virus isolates were grouped into either cluster 1 or cluster 2, which likely originated in 1997 and 2006, respectively. Cluster 1 viruses were found in all provinces examined in this study and had a mutation rate of 1.90 × 10−3 per site per year. The effective number of infections remained constant until 2007, and along with the introduction of rubella vaccine into the national immunization program, although the circulation of cluster 1 viruses has not been interrupted, some viral lineages have disappeared, and the epidemic started a decline that led to a decrease in the effective population size. Cluster 2 viruses were found only in Hainan Province, likely because of importation.
Exponential apparent diffusion coefficient (EADC) is an indicator of diffusion-weighted imaging (DWI) and reflects the pathological changes of tissues quantitatively. However, no study has been investigated in the space-occupying kidney disease using EADC values. This study aims to evaluate the diagnostic role of EADC values at a high magnetic field strength (3.0 T) in kidney neoplastic lesions, compared with that of the ADC values. Ninety patients with suspected renal tumors (including 101 suspected renal lesions) and 20 healthy volunteers were performed MRI scanning. Diffusion-weighted imaging was performed with a single-shot spin-echo echo-planar imaging (SE-EPI) sequence at a diffusion gradient of b = 500 s/mm2. We found renal cell carcinoma (RCC) can be distinguished from angiomyolipoma, and clear cell carcinoma can be distinguished from non-clear cell carcinoma by EADC value. There was significant difference in overall EADC values between renal cell carcinoma (0.150 ± 0.059) and angiomyolipoma (0.270 ± 0.108) when b value was 500 s/mm2. When receiver operating characteristic (ROC) was higher than 0.192, the sensitivity and specificity of EADC value of renal cell carcinoma were 84.6 and 81.1 %, respectively. In conclusion, EADC map shows the internal structure of the kidney tumor more intuitively than the ADC map dose, and is also in line with the observation habits of the clinicians. EADC can be used as an effective imaging method for tumor diagnosis.
This study is based on an unique dataset of birth records of 11,693 persons born in Beijing, China from 1921–54 and from clinical examinations (May 2003–April 2005) of 2,085 tracked surviving cohort members (aged 50–82) to diagnose seven major chronic diseases. Data were analyzed using the extended Fixed Attribute Dynamics (FAD) method and multivariate regressions. The results of our FAD analysis have shown that, as compared to the persons with low birth weight (<2,500 g), the probability of survival from age 0 to ages 50–56, 63–67 and 68–82 for persons with higher birth weight (2,500–2,999 g, 3,000–3499 g, or ≥3,500 g) was 16–31, 62–104, and 52–108% higher respectively. These estimates display similar patterns for both genders, with somewhat stronger effects in men. Of the 27 estimates of the Odds Ratio of Survival in the FAD analysis, 21 are statistically significant. Controlling for 14 confounding factors, multivariate binary logistic regressions have demonstrated that the risk of having cardiovascular disease and diabetes is negatively associated birth weight; ordinal logit regressions have shown that the number of major chronic diseases at ages 50–82 is significantly associated with birth weight: the lower the birth weight, the higher the risk of having more chronic diseases. We conclude that low birth weight is negatively associated with 1) long-term survival probability from age 0 to ages 50–82, and 2) overall health at middle and old ages. Further research is needed to understand the mechanisms of these effects.
Birth weight; Fetal growth; long-term survival; Chronic diseases; health; China
To determine if the utilization of injectable chemically-modified hyaluronan (HA) derivative at the time of intentional vocal fold resection may facilitate wound repair and preserve the unique viscoelastic properties of the extracellular matrix and lamina propria 6 months after treatment.
Prospective, controlled animal study.
Twelve rabbit vocal folds were biopsied bilaterally, and the left side of vocal fold was treated with Extracel, an injectable, chemically-modified HA derivative, and the right side of vocal fold was injected with saline as control at the time of resection. Animals were sacrificed six months after biopsy and injection. Outcomes measured include transcription levels for procollagen, fibronectin, fibromodulin, TGF-β1, hyaluronan synthase and hyaluronidase and tissue biomechanics -- viscosity and elasticity.
Extracel treated vocal folds were found to have significantly less fibrosis than saline treated controls. Extracel treated vocal folds had significantly improved biomechanical properties of elasticity and viscosity. Significantly decreased levels of fibronectin, fibromodulin, TGF-β1, procollagen I and hyaluronan synthase were measured.
Prophylactic in vivo manipulation of the extracellular matrix with an injectable HA hydrogel appears to induce vocal fold tissue regeneration to yield improved tissue composition and biomechanical properties at 6 months.
vocal folds; scarring; prophylactic; hyaluronic acid hydrogels; tissue engineering
We have previously shown that transplantation of immature DCX+/NeuN+/Prox1+ neurons (found in the neonatal DG), but not undifferentiated neuronal progenitor cells (NPCs) from ventral subventricular zone (SVZ), results in neuronal maturation in vivo within the dentate niche. Here we investigated whether we could enhance the integration of SVZ NPCs by forced expression of the proneural gene Neurogenin 2 (NEUROG2). NPCs cultured from neonatal GFP-transgenic rat SVZ for 7 days in a non-differentiating medium were transduced with a retrovirus encoding NEUROG2 and DsRed or the DsRed reporter gene alone (control). By 3 days post-transduction, the NEUROG2-transduced cells maintained in culture contained mostly immature neurons (91% DCX+; 76% NeuN+), whereas the control virus-transduced cells remained largely undifferentiated (30% DCX+; <1% NeuN+). At 6 weeks following transplantation into the DG of adult male rats, there were no neurons among the transplanted cells treated with the control virus but the majority of the NEUROG2-transduced DsRed+ SVZ cells became mature neurons (92% NeuN+; DCX-negative). Although the NEUROG2-transduced SVZ cells did not express the dentate granule neuron marker Prox1, most of the NEUROG2-transduced SVZ cells (78%) expressed the glutamatergic marker Tbr1, suggesting the acquisition of a glutamatergic phenotype. Moreover, some neurons extended dendrites into the molecular layer, grew axons containing Ankyrin G+ axonal initial segments, and projected into the CA3 region, thus resembling mature DG granule neurons. A proportion of NEUROG2 transduced cells also expressed c-Fos and P-CREB, two markers of neuronal activation. We conclude that NEUROG2-transduction is sufficient to promote neuronal maturation and integration of transplanted NPCs from SVZ into the DG.
Various α2,3 subtype selective partial GABA-A agonists are in development to treat anxiety disorders. These compounds are expected to be anxiolytic with fewer undesirable side effects, compared to nonselective GABA-A agonists like benzodiazepines. Several α2,3 subtype selective and nonselective GABA-A agonists have been examined in healthy volunteers, using a battery addressing different brain domains. Data from five placebo-controlled double-blind studies were pooled. Lorazepam 2 mg was the comparator in three studies. Three α2,3-selective GABAA agonists (i.e., TPA023, TPACMP2, SL65.1498), one α1-selective GABAA agonists (zolpidem), and another full agonist (alprazolam) were examined. Pharmacological selectivity was assessed by determination of regression lines for the change from baseline of saccadic-peak-velocity- (ΔSPV-) relative effect, relative to changes in different pharmacodynamic endpoints (ΔPD). SPV was chosen for its sensitivity to the anxiolysis of benzodiazepines. Slopes of the ΔSPV-ΔPD relations were consistently lower with the α2,3 selective GABA-A agonists than with lorazepam, indicating that their PD effects are less than their SPV-effects. The ΔSPV-ΔPD relations of lorazepam were comparable to alprazolam. Zolpidem showed relatively higher impairments in ΔPD relative to ΔSPV, but did not significantly differ from lorazepam. These PD results support the pharmacological selectivity of the α2,3-selective GABA-A agonists, implying an improved therapeutic window.
We developed a method to fabricate a periodic array of three-dimensional crescent-like holes (3DCLH) via an inverted hemispherical colloidal lithography. It is found that there exists an extraordinary optical transmission in this non-planar perforated periodic array of 3DCLH when the electric field of the incident light is perpendicular to the cross-line of the crescent-like hole. This extraordinary optical peak is insensitive with the incident angles and sensitive with the angle between the electric field of the incident light to the cross-line of the 3DCLH. Numerical simulation based on finite-difference time-domain method reveals that this peak is caused by an asymmetric localized surface plasmon resonance. This structure might be useful for the optical sensing and optical-integrated circuits.
Surface plasmon polariton; Localized surface plasmon; Extraordinary optical transmission; Colloidal lithography
The role of myofibroblasts in vocal fold scarring has not been extensively studied partly due to a lack of a robust in vitro model. The objective of this investigation was to develop and characterize a myofibroblast in vitro model that could be utilized to investigate the molecular mechanism of myofibroblast differentiation and function in injured vocal fold tissue. Differentiation of human primary vocal fold fibroblasts (hVFF) to myofibroblasts was stimulated using 5, 10, or 20 ng/ml of recombinant transforming growth factor beta-1 (TGF-β1). Cultures were analyzed using immunofluorescence and western blotting, with an anti-alpha smooth muscle actin (α-SMA) antibody as a myofibroblast marker. Normal rabbit vocal folds were treated with 10 ng/ml of TGF-β1 for 7 days for in vivo corroboration. The effects of interleukin-6 (IL-6) and hepatocyte growth factor (HGF) on myofibroblast differentiation were studied using western blots. hVFF demonstrated positive α-SMA labeling in 10 and 20 ng/ml TGF-β1 stimulated cells indicating that hVFFs were capable of differentiation to myofibroblasts. TGF- β1 induced the largest increase in α-SMA at 10-ng/ml on day 5 of treatment. HGF and IL6 suppressed the expression of TGF-β1 induced α–SMA. Our work characterizes a useful in vitro model of TGF-β1 mediated vocal fibroblast-myofibroblast differentiation. The extent of differentiation appears to be attenuated by HGF suggesting a potential mechanism to support prior work indicating that HGF plays a protective role from scar formation in vocal fold injuries. Paradoxically, IL-6 which has been shown to play a profibrotic role in dermal studies also attenuated the TGF-β1 response.
fibroblasts; TGF-β1; α-SMA; vocal fold; myofibroblasts; wound healing
Tumor necrosis factor-alpha (TNF-α) is an inflammatory cytokine and apoptotic molecule that appears to be a mediator in inflammation and fibrosis. The objective of this investigation was to examine the effects of TNF-α on three-dimensional (3D) Carbylan-GSX in vitro cultured human vocal fold fibroblasts (hVFF), to provide insight into the mechanism responsible for improved vocal fold wound healing that has been previous reported with Carbylan-GSX treatment.
In vitro cell culture.
hVFF were cultured in three dimensional (3D) Carbylan-GSX and on polystyrene with different dosages of TNF-α (0, 0.1, 1, 10 and 100 ng/ml) with and without 10% fetal bovine serum (FBS). hVFF response to TNF-α was characterized by morphology, proliferation rates and gene transcript levels for MMP1, MMP2, TIMP3, collagen I, collagen III, fibronectin and TNF-α receptor.
In 3D Carbylan-GSX, TNF-α inhibited hVFF proliferation in a dose-dependent manner, TNF-α (0.1–100ng/ml) was shown to significantly down-regulate TIMP3 and ECM related mRNA transcript levels for collagen III and fibronectin, and to up-regulate MMP1 and MMP2 expression, resulting in increased MMP/TIMP3 ratios. TNF-α receptor expression was significant up-regulated in Carbylan-GSX compared to control polystyrene. Responses were more marked in 10%FBS culture.
After vocal fold injury, locally injected Carbylan-GSX can enhanced the role of TNF-α in remodeling the lamina propria layer of vocal fold, accelerating wound healing, suggesting Carbylan-GSX as a potential new therapeutic approach that may lead to better treatment of vocal fold wound healing.
human vocal fold fibroblasts; Carbylan-GXS; three-dimensional (3D) cell culture; TNF-α; wound healing; fibrosis
In order to promote wound repair and induce tissue regeneration, an engineered hyaluronan (HA) hydrogel – Carbylan GSX, which contains di(thiopropionyl) bishydrazide-modified hyaluronic acid (HA-DTPH), di(thiopropionyl) bishydrazide-modified gelatin (Gtn-DTPH) and polyethylene glycol diacrylate (PEGDA), has been developed for extracellular matrix (ECM) defects of the superficial and middle layers of the lamina propria. The purpose of this study was to evaluate the biocompatibility of Carbylan GSX in a previously established immortalized human vocal fold fibroblast (hVFF) cell line prior to human clinical trials. Immortalized hVFF proliferation, viability, apoptosis and transcript analysis for both ECM constituents and inflammatory markers were measured for two-dimensional and three-dimensional culture conditions. There were no significant differences in morphology, cell marker protein expression, proliferation, viability and apoptosis of hVFF cultured with Carbylan GSX compared to Matrigel, a commercial 3D control, after one week. Gene expression levels for fibromodulin, TGF-β1, and TNF-α were similar between Carbylan GSX and Matrigel. Fibronectin, hyaluronidase 1 and COX2 expression levels were induced by Carbylan GSX; whereas IL6, IL8. COL1 and hyaluronic acid synthase 3 expression levels were decreased by Carbylan GSX. This investigation demonstrates that Carbylan GSX may serve as a natural biomaterial for tissue engineering of human vocal folds.
human vocal fold fibroblasts; three-dimensional culture; extracellular matrix; HA hydrogel (Carbylan GSX); cell proliferation; cell viability; gene expression
Streptococcus thermophilus strain ND03 is a Chinese commercial dairy starter used for the manufacture of yogurt. It was isolated from naturally fermented yak milk in Qinghai, China. We present here the complete genome sequence of ND03 and compare it to three other published genomes of Streptococcus thermophilus strains.
Lactobacillus delbrueckii subsp. bulgaricus strain ND02 is a Chinese commercial dairy starter used for the manufacture of yoghurt. It was isolated from naturally fermented yak milk in Qinghai, China. Here, we report the main genome features of ND02 and several differences with two other published genomes of Lactobacillus delbrueckii subsp. bulgaricus strains.
Lactobacillus casei Zhang is a new probiotic bacterium isolated from koumiss collected in Inner Mongolia, China. Here, we report the main genome features of L. casei Zhang and the identification of several predicted proteins implicated in interactions with the host.
Cytokine-induced killer (CIK) cell therapy, an adoptive T-cell immunotherapy, has been reported to be a safe and effective mode of treatment for patients with metastatic diseases, lymphoma and acute leukaemia. To investigate the clinical efficacy of cytokine-induced killer cells for the treatment of refractory lymphoma, the present clinical study was conducted. A total of 8 male patients with a mean age of 41 years (range 22–65) who were pathologically diagnosed with malignant lymphoma (Hodgkin's disease, 2 and non-Hodgkin's lymphoma, 6) were enrolled. CIK cells were expanded by priming with IFN-γ, monoclonal antibody (mAb) to CD3 and IL-1α, followed by the addition of IL-2 the following day using peripheral blood mononuclear cells (PBMCs) of the 8 male patients. The CIK cells were then transfused back to the patients as treatment. On day 13, the CIK cell count reached 7–18×1019 (mean, 12.7×109), a 44- to 140-fold increase (mean, 98-fold). The average percentage of cells expressing CD3+, CD4+, CD8+ and CD3+CD56+ were also increased from 50.9±3.5, 29.9±1.7, 41.3±3.2, 1.6±0.2% to 90.2±1.6, 40.6±5.5, 52.8±4.9 and 33.1±4.0%, respectively. Patients showed measurable radiographic tumor reduction, increased T-cell subset levels, and relief of symptoms after treatment. No severe toxicity or side effects were reported. CIK cells developed by this culture method have a high in vitro proliferation rate and tumor-killing capacity. In conclusion, CIK cell treatment of patients with malignant lymphoma achieves effective clinical responses, causing few side effects.
cytokine-induced killer cells; malignant lymphoma; adoptive immune therapy
To determine the effectiveness of bone marrow mesenchymal stem cell (BM-MSC) transplantation in isolation or within a synthetic extracellular matrix (sECM) for tissue regeneration of the scarred vocal fold lamina propria.
In vitro stability and compatibility of mouse BM-MSC embedded in sECM was assessed by flow cytometry detection of BM-MSC marker expression and proliferation. Eighteen rats were subjected to vocal fold injury bilaterally, followed by one month post-treatment with unilateral injections of saline or sECM hydrogel (Extracel), GFP-mouse BM-MSC or BM-MSC suspended in sECM. Outcomes measured one month after treatment included procollagen-III, fibronectin, hyaluronan synthase-III (HAS3), hyaluronidase (HYAL3), smooth muscle actin (SMA) and transforming growth factor-beta 1(TGF-β1) mRNA expression. The persistence of GFP BM-MSC, proliferation, apoptosis and myofibroblast differentiation was assessed by immunofluorescence.
BM-MSC grown in vitro within sECM express Sca-1, are positive for hyaluronan receptor CD44 and continue to proliferate. In the in vivo study, groups injected with BM-MSC had detectable GFP-labeled BM-MSC remaining, showed proliferation and low apoptotic or myofibroblast markers compared to the contralateral side. Embedded BM-MSC in sECM group exhibited increased levels of procollagen III, fibronectin and TGF-β1. BM-MSC within sECM downregulated the expression of SMA compared to BM-MSC alone, exhibited upregulation of HYAL3 and no change in HAS3 compared to saline.
Treatment of vocal fold scarring with BM-MSC injected in a sECM displayed the most favorable outcomes in ECM production, hyaluronan metabolism, myofibroblast differentiation and production of TGF-β1. Furthermore, the combined treatment had no detectable cytotoxicity and preserved local cell proliferation.
Mesenchymal stem cells; tissue engineering; hydrogel; extracellular matrix; myofibroblast differentiation
The success of transplants of neural tissue into the adult dentate gyrus in generating mature neurons is highly variable. Here we address the roles of the origin of the tissue and its pre-implantation preparation, and show that both are critical. We transplanted neonatal cultured or primary rat cells from either the ventral subventricular zone (vSVZ) or the dentate gyrus (DG) into the adult rat DG. Only primary DG cells robustly generated DG neurons (80% NeuN and Prox1-positive cells at 6 weeks), substantially repaired the damaged DG, and formed glutamatergic projections to the target CA3 region. Cultured DG cells expanded for 7 days showed limited neuronal differentiation after transplantation (10% NeuN and Prox1-positive cells) whereas cultured or primary vSVZ cells failed to make any Prox1-positive DG granular neurons. We found that a specific population of postmitotic young neurons (triple doublecortin/NeuN/Prox1-positive) were particularly abundant in primary DG cells, but were markedly reduced in the cultured DG cells and were absent in the cultured and primary vSVZ cells. Labelling of primary DG cells with the mitotic marker BrdU suggested that postmitotic young neurons are the source of the transplanted mature neurons in-vivo. We conclude that both the origin and pre-transplantation history of donor cells are key factors that determine the outcome of transplantation. These findings may be of therapeutic interest for cell replacement therapy in treating the damaged hippocampus.
Bifidobacterium animalis subsp. lactis strain V9 is a Chinese commercial bifidobacteria with several probiotic functions. It was isolated from a healthy Mongolian child in China. We present here the complete genome sequence of V9 and compare it to 3 other published genome sequences of B. animalis subsp. lactis strains. The result indicates the lack of polymorphism among strains of this subspecies from different continents.
The incidence of rubella cases in China from 1991 to 2007 was reviewed, and the nucleotide sequences from 123 rubella viruses collected during 1999 to 2007 and 4 viral sequences previously reported from 1979 to 1984 were phylogenetically analyzed. Rubella vaccination was not included in national immunization programs in China before 2007. Changes in endemic viruses were compared with incidences of rubella epidemics. The results showed that rubella epidemics occur approximately every 6 to 8 years (1993/1994, 2001, and 2007), and a shift of disease burden to susceptible young adults was observed. The Chinese rubella virus sequences were categorized into 5 of the 13 rubella virus genotypes, 1a, 1E, 1F, 2A, and 2B; cocirculations of these different genotypes were found in China. In Anhui province, a shift in the predominant genotype from 1F and 2B to 1E coincided with the 2001 rubella epidemic. This shift may have occurred throughout China during 2001 to 2007. This study investigated the genotype distribution of rubella viruses in China over a 28-year period to establish an important genetic baseline in China during its prevaccination era.
Diffuse large B-cell lymphoma (DLBCL) represents a clinically heterogeneous disease. Models based on immunohistochemistry predict clinical outcome. These include subdivision into germinal center (GC) versus non-GC subtypes; proliferation index (measured by expression of Ki-67), and expression of BCL-2, FOXP1, or B-lymphocyte-induced maturation protein (Blimp-1)/PRDM1. We sought to determine whether immunohistochemical analyses of biopsies from patients with DLBCL having HIV infection are similarly relevant for prognosis.
Patients and Methods
We examined 81 DLBCLs from patients with AIDS in AMC010 (cyclophosphamide, doxorubicin, vincristine, and prednisone [CHOP] v CHOP-rituximab) and AMC034 (etoposide, doxorubicin, vincristine, prednisone, and dose-adjusted cyclophosphamide plus rituximab concurrent v sequential) clinical trials and compared the immunophenotype with survival data, Epstein-Barr virus (EBV) positivity, and CD4 counts.
The GC and non-GC subtypes of DLBCL did not differ significantly with respect to overall survival or CD4 count at cancer presentation. EBV could be found in both subtypes of DLBCL, although less frequently in the GC subtype, and did not affect survival. Expression of FOXP1, Blimp-1/PRDM1, or BCL-2 was not correlated with the outcome in patients with AIDS-related DLBCL.
These data indicate that with current treatment strategies for lymphoma and control of HIV infection, commonly used immunohistochemical markers may not be clinically relevant in HIV-infected patients with DLBCL. The only predictive immunohistochemical marker was found to be Ki-67, where a higher proliferation index was associated with better survival, suggesting a better response to therapy in patients whose tumors had higher proliferation rates.