Electrospun biocomposite nanofibrous scaffolds for neural tissue engineering

Bridging of nerve gaps after injury is a major problem in peripheral nerve regeneration. Considering the potential application of a bio-artificial nerve guide material, polycaprolactone (PCL)/chitosan nanofibrous scaffolds was designed and evaluated in vitro using rat Schwann cells (RT4-D6P2T) for nerve tissue engineering. PCL, chitosan, and PCL/chitosan nanofibers with average fiber diameters of 630, 450, and 190 nm, respectively, were fabricated using an electrospinning process. The surface chemistry of the fabricated nanofibers was determined using Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy. Simple blending of PCL with chitosan proved an easy and efficient method for fabricating PCL/chitosan nanofibrous scaffolds, whose surface characteristics proved more hydrophilic than PCL nanofibers. Evaluation of mechanical properties showed that the Young's modulus and strain at break of the electrospun PCL/chitosan nanofibers were better than those of the chitosan nanofibers. Results of cell proliferation studies on nanofibrous scaffolds using 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium assay showed 48% more cell proliferation on PCL/chitosan scaffolds than on PCL scaffolds after 8 days of culture. PCL/chitosan scaffolds showed better cell proliferation than PCL scaffolds and maintained their characteristic cell morphology, with spreading bipolar elongations to the nanofibrous substrates. This electrospun nanofibrous matrix thus proved of specific interest in tissue engineering for peripheral nerve regeneration.     

Priming microenvironments dictate cytokine requirements for T helper 17 cell lineage commitment

Activation of pattern recognition receptors on dendritic cells (DCs) and macrophages leads to secretion of cytokines that control differentiation of CD4(+) T cells. The current understanding is that interleukin-6 (IL-6) in combination with transforming growth factor-β (TGF-β) leads to generation of T helper 17 (Th17) lineage cells. Here, we have discovered that the cytokine requirements for Th17 cell polarization depend on the site of priming. Although IL-6 played a critical role in Th17 cell lineage priming in the skin and mucosal tissues, it was not required for Th17 cell priming in the spleen. In contrast, IL-1 played an irreplaceable role for priming of Th17 lineage cells in all tissues. Importantly, we have demonstrated that IL-6-independent and -dependent pathways of Th17 cell differentiation are guided by DCs residing in various tissues. These results reveal fundamental differences by which the systemic, mucosal, and cutaneous immune systems guide Th17 cell lineage commitment.     

Guided orientation of cardiomyocytes on electrospun aligned nanofibers for cardiac tissue engineering

Cardiac tissue engineering (TE) is one of the most promising strategies to reconstruct the infarct myocardium and the major challenge involves producing a bioactive scaffold with anisotropic properties that assist in cell guidance to mimic the heart tissue. In this study, random and aligned poly(ε-caprolactone)/gelatin (PG) composite nanofibrous scaffolds were electrospun to structurally mimic the oriented extracellular matrix (ECM). Morphological, chemical and mechanical properties of the electrospun PG nanofibers were evaluated by scanning electron microscopy (SEM), water contact angle, attenuated total reflectance Fourier transform infrared spectroscopy and tensile measurements. Results indicated that PG nanofibrous scaffolds possessed smaller fiber diameters (239 ± 37 nm for random fibers and 269 ± 33 nm for aligned fibers), increased hydrophilicity, and lower stiffness compared to electrospun PCL nanofibers. The aligned PG nanofibers showed anisotropic wetting characteristics and mechanical properties, which closely match the requirements of native cardiac anisotropy. Rabbit cardiomyocytes were cultured on electrospun random and aligned nanofibers to assess the biocompatibility of scaffolds, together with its potential for cell guidance. The SEM and immunocytochemical analysis showed that the aligned PG scaffold greatly promoted cell attachment and alignment because of the biological components and ordered topography of the scaffolds. Moreover, we concluded that the aligned PG nanofibrous scaffolds could be more promising substrates suitable for the regeneration of infarct myocardium and other cardiac defects.     

CALLA negative precursor B lymphoblastic leukemia with MLL gene translocation and an unusual FISH signal pattern

Rearrangements of the mixed lineage leukemia (MLL) gene at 11q23 commonly occur in infants with CALLA negative B lymphoblastic leukemia (B-ALL). Most often, these are detected by conventional karyotyping; however, fluorescent in-situ hybridization (FISH) with the help of a dual-color break-apart probe is used to identify cryptic translocations. When there is an MLL gene translocation, the usual FISH signal pattern is 1 red-1 green-1 yellow fusion signal pattern. We present a case of an infant with CALLA negative precursor B-ALL with a characteristic translocation t(4;11) (q21;q23), however, with an unusual MLL FISH signal pattern.     

Biocompatibility evaluation of emulsion electrospun nanofibers using osteoblasts for bone tissue engineering

Emulsion electrospinning is an advanced technique to fabricate core-shell structured nanofibrous scaffolds, with great potential for drug encapsulation. Incorporation of dual factors hydroxyapatite (HA) and laminin, respectively, within the shell and core of nanofibers through emulsion electrospinning might be of advantageous in supporting the adhesion, proliferation, and maturation of cells instead of single factor-encapsulated nanofibers. We fabricated poly(L-lactic acid-co-?-caprolactone) (PLCL)/hydroxyapaptite (PLCL/HA), PLCL/laminin (PLCL/Lam), and PLCL/hydroxyapatite/laminin (PLCL/HA/Lam) scaffolds with fiber diameter of 388?±?35, 388?±?81, and 379?±?57?nm, respectively, by emulsion electrospinning. The elastic modulus of the prepared scaffolds ranged from 22.7–37.0?MPa. The osteoblast proliferation on PLCL/HA/Lam scaffolds, determined on day 21, was found 10.4% and 12.0% higher than the cell proliferation on PLCL/Lam or PLCL/HA scaffold, respectively. Cell maturation determined on day 14, by alkaline phosphatase (ALP) activity, was significantly higher on PLCL/HA/Lam scaffolds than the ALP activity on PLCL/HA and PLCL/Lam scaffolds (p???0.05). Results of the energy dispersive X-ray studies carried out on day 28 also showed higher calcium deposition by cells seeded on PLCL/HA/Lam scaffolds. Osteoblasts were found to adhere, proliferate, and mature actively on PLCL/HA/Lam nanofibers with enhanced cell proliferation, ALP activity, bone protein expression, and mineral deposition. Based on the results, we can conclude that laminin and HA individually played roles in osteoblast proliferation and maturation, and the synergistic function of both factors within the novel emulsion electrospun PLCL/HA/Lam nanofibers enhanced the functionality of osteoblasts, confirming their potential application in bone tissue regeneration.     

Mutation and Methylation Analysis of the Chromodomain-Helicase-DNA Binding 5 Gene in Ovarian Cancer

Chromodomain, helicase, DNA binding 5 (CHD5) is a member of a subclass of the chromatin remodeling Swi/Snf proteins and has recently been proposed as a tumor suppressor in a diverse range of human cancers. We analyzed all 41 coding exons of CHD5 for somatic mutations in 123 primary ovarian cancers as well as 60 primary breast cancers using high-resolution melt analysis. We also examined methylation of the CHD5 promoter in 48 ovarian cancer samples by methylation-specific single-stranded conformation polymorphism and bisulfite sequencing. In contrast to previous studies, no mutations were identified in the breast cancers, but somatic heterozygous missense mutations were identified in 3 of 123 ovarian cancers. We identified promoter methylation in 3 of 45 samples with normal CHD5 and in 2 of 3 samples with CHD5 mutation, suggesting these tumors may have biallelic inactivation of CHD5. Hemizygous copy number loss at CHD5 occurred in 6 of 85 samples as assessed by single nucleotide polymorphism array. Tumors with CHD5 mutation or methylation were more likely to have mutation of KRAS or BRAF (P = .04). The aggregate frequency of CHD5 haploinsufficiency or inactivation is 16.2% in ovarian cancer. Thus, CHD5 may play a role as a tumor suppressor gene in ovarian cancer; however, it is likely that there is another target of the frequent copy number neutral loss of heterozygosity observed at 1p36.     

Poststreptococcal reactive arthritis in adults: a case series

OBJECTIVE: To guide primary care physicians regarding the diagnosis and treatment of poststreptococcal reactive arthritis (PSReA) in adults. PATIENTS AND METHODS: We retrospectively reviewed an indexed database of all patients evaluated or hospitalized between 1976 and 1998 at Mayo Clinic Rochester and identified 35 patients with the diagnosis of reactive streptococcal arthritis, arthralgia, or arthritides. Twenty-nine patients with the diagnosis of acute rheumatic fever (ARF), septic streptococcal arthritis, or nonspecific reactive arthritis were excluded. RESULTS: PSReA was confirmed in 6 adults (3 women, 3 men; age range, 25-66 years). All patients were symptomatic with polyarthritis and oligoarthritis disproportionate to the objective findings on physical examination. Although all patients had negative throat cultures at the onset of arthritis, increased titers of anti-DNase B and antistreptolysin O confirmed recent streptococcal infection. Antecedent events included pharyngitis in 3 patients (who had received a minimum of a 10-day course of penicillin) and toxic shock syndrome in 1 patient. The latency of onset of arthritis ranged from 4 days to 6 weeks. The arthritic symptoms had a protracted course beyond the typical maximum of 3 weeks described for ARF. Treatment with aspirin did not provide symptomatic relief in any of the patients, whereas the response to therapy with nonsteroidal anti-inflammatory drugs (NSAIDs) was at least partial in all cases. Symptomatic relief occurred in 1 patient who received indomethacin and in 1 patient treated with prednisone. Penicillin prophylaxis was recommended in 1 patient. CONCLUSION: PSReA should be included in the differential diagnosis of all adult patients presenting with arthritis. Treatment strategies include aspirin, other NSAIDs, and corticosteroids. In adult patients with PSReA, there is no evidence to support the use of penicillin prophylaxis at this time.     

Pulmonary tuberculosis in ahmedabad: Epidemiology,diagnosis and short course chemotherapy

Five hundred children below the age of 12 years suffering from lung tuberculosis viz., primary complex (PC) or progressive primary complex (PPC) were studied. Diagnosis was based on Kenneth Jones criteria; selected cases having score of 5 or more. One hundred and eighty cases of PC were given A-1 (6 RH) regimen, while 312 cases of PPC were given A-2 (2SHRZ/4 RH) or A-3 (2 SRH/4 RH) or A-4 (2 RHE/4 RH) regimen. Follow-up was done for 6 months after completing the treatment to observe the relapse rate. In cases of PC, 6 RH regimen appeared adequate and cheaper with no relapse rate. In cases of PPC with short course chemotherapy, compliance of patients had been very good. Relapse rate was upto 13% which is acceptable. Drug toxicity was very low.     

Role of combined therapy in advanced carcinoma of the larynx and hypopharynx

The present prospective preliminary study is based on treatment of stage III and IV cancers of the larynx and hypopharynx with a planned combined regimen of radiotherapy and radical surgery. There is a reduction in the failure rate in these cases as compared to those treated in this centre with a single modality of therapy like radiotherapy or surgery alone. 69% of the patients thus treated are surviving disease-free for follow ups varying 18–30 months with a median follow up of 24 months following combined treatment.     

Alterations in membrane-bound and cytoplasmic K-ras protein levels in mouse lung induced by treatment with lovastatin,cholestyramine, or niacin: effects are highly mouse strain dependent

Agents that either increase (cholestyramine, CS) or decrease (lovastatin, Lov) de novo peripheral cholesterol synthesis may increase (CS) or decrease (Lov) ras protein membrane localization by altering protein prenylation, and potentially have pro- or anti-carcinogenic effects. Male A/J, Swiss, and C57/BL6 mice were treated with 2 or 4% CS, 1% dietary niacin, or 25mg/kg of Lov three times per week (Lov-3X) or five times per week (Lov-5X). After 3 weeks, serum cholesterol and triglycerides were determined enzymatically. Membrane and cytoplasmic K-ras proteins in lung were determined by immunoprecipitation followed by western blotting with a K-ras specific antibody. Results confirmed the hypothesis only in isolated instances. A/J mice had a significant 30% increase in cytoplasmic K-ras and a 40% decrease in membrane K-ras from Lov treatment, as predicted. C57/BL6 mice had a significant 77% increase in membrane K-ras, as expected from CS feeding. At variance with the hypothesis, Swiss mice had increased levels (3-28%) of membrane K-ras with all treatments (including Lov), and C57/BL6 mice treated with Lov had a 58-78% increase in cytoplasmic K-ras without any reduction in the levels of membrane K-ras. Niacin, predicted to have no effect on ras membrane localization, decreased cytoplasmic K-ras in A/J mice, increased both membrane and cytoplasmic K-ras in Swiss mice, and had no effect in C57/BL6 mice. Results may have differed from those predicted because of strain-dependent differences in response to the cholesterol-lowering agents. A difference in response among the mouse strains suggests that individual genetic differences may alter the effect of hypocholesterolemic agents on K-ras membrane localization, and potentially the risk of ras-dependent cancer.