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Several functional analyses are conducted to explain the ability of the deletions and point mutations of LRRC50 in causing the primary ciliary dyskinesia (PCD). The outer and the inner dynein arm defects are shown to be the main defects that lead to the onset of the disorder.

This study group has previously identified IL-9–producing mucosal mast cell (MMC9) as the primary source of IL-9 to drive intestinal mastocytosis and experimental IgE-mediated food allergy. However, the molecular mechanisms that regulate the expansion of MMC9s remain unknown. This study hypothesized that IL-4 regulates MMC9 development and MMC9-dependent experimental IgE-mediated food allergy. An epicutaneous sensitization model was used and bone marrow reconstitution experiments were performed to test the requirement of IL-4 receptor α (IL-4Rα) signaling on MMC9s in experimental IgE-mediated food allergy. Flow cytometric, bulk, and single-cell RNA-sequencing analyses on small intestine (SI) MMC9s were performed to illuminate MMC9 transcriptional signature and the effect of IL-4Rα signaling on MMC9 function. A bone marrow–derived MMC9 culture system was used to define IL-4–BATF signaling in MMC9 development. Epicutaneous sensitization– and bone marrow reconstitution–based models of IgE-mediated food allergy revealed an IL-4 signaling-dependent cell-intrinsic effect on SI MMC9 accumulation and food allergy severity. RNA-sequencing analysis of SI-MMC9s identified 410 gene transcripts reciprocally regulated by IL-4 signaling, including Il9 and Batf. In silico analyses identified a 3491-gene MMC9 transcriptional signature and identified 2 transcriptionally distinct SI MMC9 populations enriched for metabolic or inflammatory programs. Employing an in vitro MMC9-culture model system showed that generation of MMC9-like cells was induced by IL-4 and this was in part dependent on BATF. IL-4Rα signaling directly modulates MMC9 function and exacerbation of experimental IgE-mediated food allergic reactions. IL-4Rα regulation of MMC9s is in part BATF-dependent and occurs via modulation of metabolic transcriptional programs. [ABSTRACT FROM AUTHOR]

Graphical abstract Abstract Bone loss around tooth extraction sites can occur, thus making future placement of dental implants difficult. Alveolar bone regeneration can be guided by the application of a nanofibrous bone graft coupled with osteoinductive proteins/peptides, following tooth loss or tooth extraction. In the present study, we demonstrate the potential of mineralized nanofiber segments coupled with calcium-binding bone morphogenetic protein 2 (BMP-2) mimicking peptides for periodontal bone regeneration. Thin electrospun nanofiber membranes of PLGA-collagen-gelatin (2:1:1 wt ratios) were mineralized in 10× modified simulated body fluid (10× mSBF) and cryocut to segments of 20 µm. For predetermined weights of the mineralized nanofiber segments, it was possible to load various amounts of heptaglutamate E7-domain-conjugated BMP-2 peptide. Mineralized short fiber grafts (2 mg), with and without E7-BMP-2 peptides, were implanted into 2 mm × 2 mm (diameter × depth) critical-sized socket defects created in rat maxillae, following extraction of the first molar teeth. A sustained release profile of E7-BMP-2 from the mineralized nanofiber segments was recorded over 4 weeks. X-ray microcomputed tomography (µ-CT) analysis of peptide-loaded nanofiber graft filled defects revealed ∼3 times greater new bone volume and bone mineral density over 4 weeks in comparison to unfilled control defects. Further, histopathology data confirmed the formation of greater new osseous tissue in the BMP2 peptide-loaded, mineralized nanofiber segment group than that of fibrous connective tissue in the unfilled defect group. Altogether, the mineralized nanofiber segments coupled with E7-BMP-2 peptides may be an effective treatment option for alveolar bone loss and defects. Statement of Significance With the high incidence of dental implants/fixtures for missing teeth, the success of the surgical procedures in restorative dentistry is dictated by the quality and quantity of the supporting alveolar bone. To address the problem of alveolar bone loss and defects due to tumor, periodontitis, or even postextraction remodeling, the present study is the first report on the application of mineralized nanofiber fragments coupled with calcium-binding osteoinductive BMP-2 peptides as a synthetic graft material for oral bone regeneration. The ease of fabrication and application of cryocut mineralized nanofiber fragments as maxillofacial bone defect fillers present a promising alternative to the current dental bone graft formulations. Furthermore, the nanofiber segments may also be utilized for several biomedical applications including hemostasis, soft tissue engineering, and wound healing. [ABSTRACT FROM AUTHOR]

Introduction Juvenile idiopathic arthritis in temporomandibular joints (TMJs) is often treated with intra-articular steroid injections, which can inhibit condylar growth. The purpose of this study was to compare simvastatin (a cholesterol-lowering drug that reduces TMJ inflammation) with the steroid triamcinolone hexacetonide in experimental TMJ arthritis. Methods Joint inflammation was induced by injecting complete Freund's adjuvant (CFA) into the TMJs of 40 growing Sprague Dawley rats; 4 other rats were left untreated. In the same intra-articular injection, one of the following was applied: (1) 0.5 mg of simvastatin in ethanol carrier, (2) ethanol carrier alone, (3) 0.15 mg of triamcinolone hexacetonide, (4) 0.5 mg of simvastatin and 0.15 mg of triamcinolone hexacetonide, or (5) nothing additional to the CFA. The animals were killed 28 days later, and their mandibles were evaluated morphometrically and with microcomputed tomography. Results The analysis showed that the TMJs subjected to CFA alone had decreased ramus height compared with those with no treatment ( P <0.05). Groups that had injections containing the steroid overall had decreases in weight, ramus height, and bone surface density when compared with the CFA-alone group ( P <0.0001). Groups that had injections containing simvastatin, however, had overall increases in weight ( P <0.0001), ramus height ( P <0.0001), condylar width ( P <0.05), condylar bone surface density ( P <0.05), and bone volume ( P <0.0001) compared with the groups receiving the steroid injections, and they were not different from the healthy (no treatment) group. Conclusions Treatment of experimentally induced arthritis in TMJs with intra-articular simvastatin preserved normal condylar bone growth. [ABSTRACT FROM AUTHOR]

Purpose: Juvenile idiopathic arthritis often affects the temporomandibular joint (TMJ), resulting in facial deformities, and intra-articular injections of anti-inflammatory steroids used in treatment may inhibit bone growth in the developing condyle. The purpose of this pilot study was to evaluate the anti-inflammatory properties of simvastatin (SIM), a bone anabolic drug, compared with the common steroid triamcinolone hexacetonide (TH) in experimental TMJ arthritis of growing rats. Methods: Joint inflammation was induced by injecting complete Freund''s adjuvant (CFA) into the TMJs of 32 growing (4-week-old) Sprague-Dawley rats while simultaneously receiving 1) ethanol drug carrier, 2) 0.1 mg of SIM, 3) 0.5 mg of SIM, or 4) 0.15 mg of TH. Six rats had no treatment to the TMJ. Animals were euthanized 28 days later, and TMJs were decalcified and stained with hematoxylin-eosin. Results: Histopathologic TMJ results showed that CFA injection along with drug carrier induced increased thickness of the articular layer on the head of the condyle and inflammation of the retrodiscal area (CFA and ethanol). Although both TH and SIM reduced the articular layer thickness, 0.5 mg of SIM was more effective at reducing subsynovial inflammation. Conclusions: Intra-articular simvastatin showed anti-inflammatory properties in this TMJ model, prompting its further study in the growing TMJ, where bone anabolic properties would be important. [Copyright &y& Elsevier]

Abstract: Single nucleotide polymorphisms (SNPs) are believed to contain relevant information and have been therefore extensively used as genetic markers in population and conservation genetics, and molecular ecology studies. This study reports on the identification of potential SNPs in a diploid European sea bass Dicentrarchus labrax genome by using reference sequences from three assembled chromosomes and mapping all WGS datasets onto them (3× Sanger, 3× 454 and 20× SOLEXA). A total of 20,779 SNPs were identified over the 1469 gene loci and intergenic space analysed. Within chromosomes the occurrence of SNPs was the lowest in exons and higher in introns and intergenic regions, which may be explained by the fact, that coding regions are under strong selective pressure to maintain their biological function. The ratio of nonsynonymous to synonymous mutations was smaller than one for all the chromosomes, suggesting that most of deleterious nonsynonymous mutations were eliminated by negative selection. SNPs were not uniformly distributed over the chromosomes. Two chromosomes exhibited large regions with extremely low SNP density, which might represent homozygous regions in the diploid genome. The results of this study show how SNP detection can take profit from sequencing a single diploid individual, but also uncover the limits of such an approach. SNPs that have been identified will support marker development for genetic linkage mapping, population genetics and aquaculture related questions in general. [ABSTRACT FROM AUTHOR]

Abstract: In the crustacean Cherax quadricarinatus, alterations of multi-transcript expression patterns between intermolt and late premolt stages were identified in the hypodermis and in the gastrolith disk via a cDNA microarray. The gastrolith disk is a specialized epithelium forming the gastroliths at premolt. The gastroliths are deposits of calcium carbonate derived from the digested cuticle contributing the mineral to the newly formed exoskeleton at postmolt. The late premolt stage was characterized by a dramatic general up-regulation of genes in the gastrolith disk. This phenomenon is explained by the gastrolith disk function rapid formation of the relatively large gastrolith during a short period of time. Besides genes of general importance for this dramatic change, three genes related to the chitin–protein–mineral structure were identified. The cDNA and the deduced protein of the novel one of them, the chitin deacetylase 1 (Cq-CDA1) was fully characterized and its resemblance to already characterized structural proteins of the gastrolith matrix was described. Cq-CDA1 characteristics strongly indicate its participation in the gastrolith construction, although its protein product was not identified yet in the gastrolith. In addition, many differentially expressed genes with unknown function were elucidated. An unexpected milder down-regulation was observed in the hypodermis. [Copyright &y& Elsevier]

Abstract: RT-PCR is the method of choice for detecting BCR–ABL in CML and ALL. The three predominant mRNA transcripts found are e1a2 (in ALL), e13a2, and e14a2 (in CML and ALL). However, a number of “atypical”BCR–ABL transcripts (e1a3, e13a3, e14a3, e19a2, e6a2, e8a2, etc.) resulting from chromosomal breakpoints outside ABL intron 1 or BCR intron 1, 13 or 14, respectively, have been reported. These atypical transcripts may escape detection when using methods that are optimized to detect just the typical ones. We present here a novel, fast, and reliable multiplex PCR for improved detection of typical and atypical BCR–ABL transcripts. [Copyright &y& Elsevier]

Purpose: The aim of this pilot study was to determine the relationship between gelatinase (MMP-9 and MMP-2) markers of soft tissue inflammation/turnover at the bone/soft tissue interface and bone turnover (osteocalcin [OC], pyridinoline cross-linked carboxyl-terminal telopeptide of type 1 collagen [ICTP], and bone fill) during healing of an alveolar bone defect. Materials and Methods: Ten subjects undergoing oral surgery had a 5 × 5-mm trephine defect created on an edentulous ridge and were sampled at the bone/soft tissue interface at baseline (prior to flap reflection), 2 weeks and 12 weeks postsurgery, using a novel bone wash device. Recovered irrigants were analyzed for MMP-9 and MMP-2 by gelatin zymography, OC and ICTP with radioimmunoassays, and albumin (ALB; to normalize markers for blood content) with a sandwich enzyme-linked immunosorbent assay. Bone fill at 12 weeks was analyzed by radiographic absorptiometry. Results: All markers of enzymatic activity and bone turnover varied significantly across time (P ≤ .03), with bone turnover markers OC and ICTP decreasing between baseline and 2 weeks, and MMP-9 and MMP-2 increased. Measures generally returned to near baseline levels after 12 weeks. MMP-9 versus MMP-2 (r = 0.97, P < .0001) and OC versus ICTP (r = 0.38, P = .048) were correlated with each other, while MMP-9 and MMP-2 were negatively correlated with ICTP (r = −0.48, P = .011 and r = −0.62, P = .006, respectively). MMP-9 was negatively correlated with subsequent bone fill (r = −0.63, P = .07). Conclusions: Bone wash sampling showed that gelatinase activity at 2 weeks following creation of an alveolar defect appeared to decrease bone turnover and eventual bone fill, suggesting benefits for anti-MMP therapy during wound healing. [Copyright &y& Elsevier]

Summary: A microbial mat from the Black Sea shelf was analyzed by a metagenomic approach. While the habitat and its microbial community are characterized by anaerobic methane oxidation, a 79kb contiguous DNA sequence obtained from the same mat provided first evidence for the concomitant presence of the capacity for anaerobic benzoate degradation. Benzoyl-CoA is one central intermediate of anaerobic aromatic degradation, among others. Within a stretch of 31kb, all genes required for the complete pathway of anaerobic benzoate degradation (catabolic island) were identified, including the four subunits of the key enzyme benzoyl-CoA reductase (bcrCBAD), which catalyzes the ATP-driven 2-electron reduction of the aromatic ring. Genes for a ketoacid:acceptor oxidoreductase (korABC) and a ferredoxin (fdx), which are required for generation of a suitable electron donor, were also detected. The majority of the identified catabolic gene products are most similar to their respective orthologs from the denitrifying freshwater bacterium Azoarcus evansii, and the genes are also similarly organized. Due to the lack of established markers, the phylogenetic affiliation of the source organism remains unclear. The presented findings indicate that the metabolic diversity of the Black Sea mat is wider than currently known and that probably other bacteria than those of the methane-oxidizing consortia contribute to aromatic degradation in this anoxic habitat. [Copyright &y& Elsevier]

The purpose of this study was to determine whether pulpal responses to cold temperatures applied to enamel, using a method that precisely controls the intensity of the cold stimulus or measures the response time, could distinguish dentin-sensitive teeth from nonsensitive teeth. Eighteen human subjects were stimulated with cold temperatures decreasing in 5°C intervals (and with tetrafluoroethane) on exposed root and enamel of a dentin-sensitive tooth and enamel of a contralateral nonsensitive tooth. Pain threshold, intensity of pain, time to pain onset, and duration of pain at baseline, 4 h, 8 h, and 1 week were measured. Responses to enamel stimulation of sensitive teeth compared with the nonsensitive teeth usually were highly correlated and not significantly different. The exception was a longer duration of pain in the dentin-sensitive teeth (4.62 ± 0.47 s) compared with nonsensitive teeth (2.92 ± 0.49 s; p = 0.016) after enamel stimulation with tetrafluoroethane. Longitudinal studies are necessary to determine whether these slight increases in pain duration indicate an increased probability of pulpal degeneration or need for dentin protection. [Copyright &y& Elsevier]

The purpose of this study was to determine the effect of one dose of a common narcotic-based pain reliever (Vicodin) on a battery of oral sensitivity tests across time in women. Fifteen Caucasian women randomly were given an oral dose of 10 mg of hydrocodone/1000 mg of acetaminophen or placebo in a double-blind, cross-over design. At baseline (before drug) and after 2, 4, and 8 h each subject was evaluated for sensitivity thresholds with four tests around an experimental tooth: (a) electric pulp tester applied to exposed root; (b) electric pulp tester on adjacent mucosa; (c) increasing probe pressure (grams) on adjacent mucosa; and (d) decreasing cold probe (°C) on the exposed root. The outcomes of all tests were not statistically different between drug and placebo treatments at any time point (p > 0.05). These results suggest that a systemic dose of hydrocodone/acetaminophen has little impact on healthy pulp or mucosa sensitivity in women as measured by common diagnostic tests. [Copyright &y& Elsevier]

Biomimetic and injectable nanofiber microspheres (NMs) could be ideal candidate for minimally invasive tissue repair. Herein, we report a facile approach to fabricate peptide-tethered NMs by combining electrospinning, electrospraying, and surface conjugation techniques. The composition and size of NMs can be tuned by varying the processing parameters during the fabrication. Further, bone morphogenic protein-2 (BMP-2) and vascular endothelial growth factor (VEGF) mimicking peptides have been successfully tethered onto poly(ε-caprolactone) (PCL):gelatin:(gelatin-methacryloyl) (GelMA)(1:0.5:0.5) NMs through photocrosslinking of the methacrylic group in GelMA and octenyl alanine (OCTAL) in the modified peptides. The BMP-2-OCTAL peptide-tethered NMs significantly promote osteogenic differentiation of bone marrow-derived stem cells (BMSCs). Moreover, human umbilical vein endothelial cells (HUVECs) seeded on VEGF mimicking peptide QK-OCTAL-tethered NMs significantly up-regulated vascular-specific proteins, leading to microvascularization. The strategy developed in this work holds great potential in developing a biomimetic and injectable carrier to efficiently direct cellular response (Osteogenesis and Angiogenesis) for tissue repair. A facile approach for the fabrication of stable peptide decorated nanofibrous microsphere for the cellular responses like osteogenesis and angiogenesis. The dual peptide decorated nanofibrous microsphere would be an appealing choice for the tissue engineering applications. Unlabelled Image [ABSTRACT FROM AUTHOR]

The Dnmt3a DNA methyltransferase has been shown to bind cooperatively to DNA and to form large multimeric protein/ DNA fibers. However, it has also been reported to methylate DNA in a processive manner, a property that is incompatible with protein/DNA fiber formation. We show here that the DNA methylation rate of Dnmt3a increases more than linearly with increasing enzyme concentration on a long DNA substrate, but not on a short 30-mer oligonucleotide substrate. We also show that addition of a catalytically inactive Dnmt3a mutant, which carries an amino acid exchange in the catalytic center, increases the DNA methylation rate by wild type Dnmt3a on the long substrate but not on the short one. In agreement with this finding, preincubation experiments indicate that stable protein/ DNA fibers are formed on the long, but not on the short substrate. In addition, methylation experiments with substrates containing one or two CpG sites did not provide evidence for a processive mechanism over a wide range of enzyme concentrations. These data clearly indicate that Dnmt3a binds to DNA in a cooperative reaction and that the formation of stable protein/ DNA fibers increases the DNA methylation rate. Fiber formation occurs at low μM concentrations of Dnmt3a, which are in the range of Dnmt3a concentrations in the nucleus of embryonic stem cells. Understanding the mechanism of Dnmt3a is of vital importance because Dnmt3a is a hotspot of somatic cancer mutations one of which has been implicated in changing Dnmt3a processivity. [ABSTRACT FROM AUTHOR]

Abstract: Summary: Antibody titers to P. gingivalis are increased in patients with rheumatoid arthritis and are associated with disease-specific autoimmunity. Background: Periodontitis (PD) has been implicated as a risk factor for rheumatoid arthritis (RA). We sought to characterize antibody titers to P. gingivalis (a pathogen in PD) in subjects with RA, PD, and in healthy controls and to examine their relationship with disease autoantibodies. Methods: P. gingivalis antibody was measured in subjects with RA (n =78), PD (n =39), and in controls (n =40). Group frequencies of bacterial titer elevations were compared using the Chi-square test and antibody titers were compared using non-parametric tests. Correlations of P. gingivalis titer with C-reactive protein (CRP), antibody to cyclic citrullinated peptide (anti-CCP), and rheumatoid factor (RF) were examined in those with RA while CRP and autoantibody concentrations were compared based on seropositivity to P. gingivalis. Results: Antibody titers to P. gingivalis were highest in PD, lowest in controls, and intermediate in RA (p =0.0003). Elevations in P. gingivalis (titer≥800) were more common in RA and PD (67% and 77%, respectively) than in controls (40%) (p =0.002). In RA, there were significant correlations with P. gingivalis titer with CRP, anti-CCP-IgM, and -IgG-2. CRP (p =0.006), anti-CCP-IgM (p =0.01) and -IgG2 (p =0.04) concentrations were higher in RA cases with P. gingivalis titers ≥800 compared to cases with titers <800. Conclusion: Antibodies to P. gingivalis are more common in RA subjects than controls, although lower than that in PD. Associations of P. gingivalis titers with RA-related autoantibody and CRP concentrations suggests that infection with this organism plays a role in disease risk and progression in RA. [Copyright &y& Elsevier]

Abstract: Objective: Osteogenic protein-1 (OP-1) has been shown to stimulate undifferentiated cells to produce mineralized tissue. Adipose tissue is a rich source of undifferentiated cells for tissue engineering purposes. The purpose of this study was to investigate the effect of OP-1 on osteogenic differentiation of adipose-derived stem cells and the production of bony tissue in vitro. Design: Adipose-derived stem cells (ADSCs) were isolated from inguinal fat pads of adult mice. Following cell expansion the cells were plated in 8-well chambered slides. The cells received one of four treatments: Group 1 cells were maintained in control medium, Group 2 cells were cultured in a common osteogenic medium, Group 3 cells were cultured in osteogenic medium supplemented with 250ng/mL of OP-1, and Group 4 cells were cultured with 250ng/mL of OP-1 added to control medium. Osteogenic differentiation of ADSCs was determined by estimating the number and size of mineralized nodules, and the amount of extracellular osteopontin secreted into cell culture medium. Mineralized nodule production was assessed at day 21 with von Kossa staining. Extracellular osteopontin release was measured after 8 and 21 days by enzyme-linked immunosorbant assay (ELISA). ANOVA/Tukey tests were used to identify differences among the four treatment groups for mineralized nodule production and osteopontin release (p ≤0.05). Results: Deposition of calcified nodules and osteopontin secretion was significantly greater for cell cultures incubated with OP-1 (p ≤0.05). At day 21, no significant differences in osteopontin secretion were noted among groups incubated with osteogenic nutrients and/or OP-1 (p >0.05), which were significantly higher than the group incubated in cell growth medium only (p ≤0.05). No significant differences in osteopontin secretion were noted between 8 and 21 days for any group (p >0.05). Linear regression analysis demonstrated a linear relationship was present between the presence of calcified nodules and the amount of osteopontin released (p ≤0.05). Conclusions: OP-1 is a powerful inducer of osteogenic differentiation of adult adipose-derived stem cells. [Copyright &y& Elsevier]

Abstract: We report the establishment and characterization, including HLA-typing, immunophenotypic and molecular cytogenetic analysis, of a novel EBV-negative cell line (BLUE-1) derived from adult relapsed sporadic Burkitt lymphoma. BLUE-1 carries the pathognomonic t(8;14)(q24;q32) effecting MYC/IgHJ fusion and a novel t(6;20)(q15;q11.2) originally present in the patient, analysis of which may facilitate identification of gene target(s) of recurrent 6q rearrangements in B-cell neoplasia. Our findings are discussed in light of the current understanding of endemic and sporadic Burkitt lymphoma. BLUE-1 grows well in culture and should be a useful lymphoma research tool. [Copyright &y& Elsevier]

The identification of genetic markers for traits of interest for aquaculture, such as growth, is an important step for the establishment of breeding programmes. As more genomic information becomes available the possibility of applying comparative genomics to identify and refine quantitative trait locus (QTLs) and potentially identify candidate genes responsible for the QTL effect may accelerate genetic improvement in established and new aquaculture species. Here we report such an approach on growth related traits in the European sea bass ( Dicentrarchus labrax ), an important species for European aquaculture. A genetic map was generated with markers targeted to previously identified QTL for growth which reduced distance and improved resolution in these regions. A total of 36 significant QTLs were identified when morphometric traits were considered individually in maternal half sibs, paternal half sibs and sib-pair analysis. Twenty seven new markers targeted to the growth QTLs, obtained by comparative mapping, reduced the average distance between markers from 23.4, 9.1, and 5.8 cM in the previous map to 3.4, 2.2, and 5.2 cM, on linkage group (LG) LG4, LG6 and LG15 respectively. Lists of genes embedded in the QTL – 591 genes in LG4, 234 genes in LG6 and 450 genes in LG15 – were obtained from the European sea bass genome. Comparative mapping revealed conserved gene synteny across teleost fishes. Functional protein association network analysis with the gene products of the 3 linkage groups revealed a large global association network including 42 gene products. Strikingly the association network was populated with genes of known biological importance for growth and body weight in terrestrial farm animals, such as elements of the signaling pathways for Jak–STAT, MAPK, adipocytokine and insulin, growth hormone, IGFI and II. This study demonstrates the feasibility of a comparative genomics combined with functional gene annotation to refine the resolution of QTL and the establishment of hypothesis to accelerate discovery of putative responsible genes. Statement of relevance This study demonstrates the feasibility of a comparative genomics approach, combined with functional annotation to refine the resolution of QTL and establishment of hypothesis to accelerate discovery of candidate genes. As production of genomic data is becoming more accessible, the implementation of this strategy will rapidly and efficiently provide the tools required for genetic selection in new candidate aquaculture species. [ABSTRACT FROM AUTHOR]

Abstract: The chromosomal translocation t(9;22)(q34;q22), with expression of the BCR-ABL1 fusion gene is the cytogenetic and molecular hallmark of chronic myeloid leukemia (CML) and a subset of acute lymphoblastic leukemia (ALL). Basically two types of BCR-ABL1 chimeric mRNA transcripts have been observed: (1) e13a2/e14a2 transcripts in CML and ALL, resulting from chromosomal breaks in the major breakpoint cluster region (M-bcr) of the BCR gene and (2) e1a2 transcripts in ALL resulting from breaks in the minor breakpoint cluster region (m-bcr) of the BCR gene. To gain a better understanding of this molecular alteration, we developed a long-distance inverse PCR (LDI PCR) method for M-bcr breakpoint identification in BCR-ABL1-positive cases and were thus able to identify the chromosomal breakpoints within the M-bcr in 62 BCR-ABL1-positive samples. The corresponding reciprocal breakpoints were identified and molecularly characterized in 45 of these cases. In 2 samples, the breaks were located 5′ to the ABL1 locus and in one case, the der(9) break was identified on 9q34.13 several hundred kB 3′ telomeric to ABL1. The analysis of breaks revealed no significant clustering and no association with repetitive elements (Alu, L1, L2) or recombination signal sequence sites. The established LDI PCR permits fast, relatively easy and unbiased identification of breakpoints in the M-bcr region of BCR and also enables the molecular analysis of more complex translocations with breakpoints outside the ABL1 gene locus or other BCR fusion genes. [Copyright &y& Elsevier]

Abstract: Resistance to treatment is a growing problem in efforts to control Old World leishmaniasis. Parasites resistant to new therapeutics such as miltefosine have not been reported from the field yet but based on experimental evidence, may appear soon. Therefore, we attempted to identify genetic markers that may correlate with miltefosine resistance. Using a functional cloning approach, we have isolated a gene from Leishmania infantum that, upon over-expression, confers protection not only against miltefosine, but also against SbIII, the active principle of anti-leishmanial antimonials. The gene encodes a very large putative polypeptide of 299kDa that shows no similarities to known proteins or functional motifs. Database mining and karyotyping experiments suggest that in L. infantum this gene is part of a 44-kbp duplicated region that is found on two separate chromosomes, CHR08 and CHR29. [Copyright &y& Elsevier]

Abstract: Systemic simvastatin is known to reduce cholesterol and stimulate modest bone formation, but local surgical placement in polylactic acid domes causes robust bone formation and local swelling. A less invasive and more flexible injection protocol was studied to evaluate the bone-inducing effects compared to surgical implantation. Bone formation rate, short- and long-term bone augmentation histology, and mechanical properties were evaluated to characterize the new bone in a rat bilateral mandible model (test and control sides in same animal). Results demonstrated that multiple (3) injections of 0.5mg simvastatin effectively reduced soft tissue swelling while preserving bone growth (60% increase of bone width at 24 days) compared to simvastatin dome placement (43% increase at 24 days). Compared to controls, bone formation rate was significantly higher on the simvastatin side, especially in the dome. Three-point bending tests revealed higher maximum force to fracture and stiffness at 24 days with simvastatin injections. Long-term evaluation showed that 55% of maximum new bone formed 24 days post-injection was retained at 90 days. [Copyright &y& Elsevier]

Abstract: An osteotropic alendronate-β-cyclodextrin conjugate (ALN-β-CD) was developed as a bone-targeting delivery system for improved treatment of skeletal diseases. The conjugate shows very strong binding to hydroxyapatite (HA, main component of the skeleton). Its ability in forming molecular inclusion complex with prostaglandin E1 (PGE1, a potent bone anabolic agent) was confirmed by phase solubility experiments and differential scanning calorimetry (DSC). In a bilateral rat mandible model, ALN-β-CD/PGE1 molecular complex was shown to stimulate strong local bone anabolic reaction. In the control study, ALN-β-CD itself was also found to be bone anabolic. To investigate this finding, other control groups were studied. The histomorphometry data suggest that ALN-β-CD itself could generate more new bone at the injection site than its complex with PGE1. Alendronate (ALN) injection could also cause new bone formation, which locates peripheral to the site of injection. PGE1, saline or ethanol injections do not have anabolic effect. These findings were also confirmed by micro-CT evaluation of mandibular bones. It is clear that the bone anabolic effect of ALN-β-CD is independent of mechanical stimuli of the periosteum or ALN injection alone. Further studies are warranted to understand the working mechanism of ALN-β-CD as a bone anabolic agent. [Copyright &y& Elsevier]

Ras-related C3 toxin substrate 1 (Rac1) is a small Rho-GTPase with important functions in fundamental cellular processes such as cytoskeleton rearrangements, signal transduction, cell cycle progression and malignant transformation. Using Rac1 primer, we identified a 5.5-kb DNA sequence on chromosome 4 (Chr. 4) in the human genome, containing the intronless protein coding sequence of Rac1. Sequence analysis revealed features of a processed pseudogene, which we named ψ1Rac1, that could be detected by Southern blot and polymerase chain reaction (PCR) on genomic DNA. A ψ1Rac1 pseudogene transcript was not detected by reverse transcription-polymerase chain reaction (RT-PCR), nor had the ψ1Rac1 promoter any transcriptional activity. In addition, three other intronless pseudogenes of Rac1 on chromosomes 4, 13 and X were identified (ψ1Rac1–ψ4Rac1) sharing an 86–96% sequence similarity with Rac1. Neither RT-PCR with pseudogene specific restriction enzymes, nor the sequencing of 130 cDNA clones from benign and malignant breast tissue and cell lines, detected the transcription of any of the Rac1 pseudogenes (ψ2Rac1–ψ4Rac1). Existence of Rac1 pseudogenes should be taken into consideration when analyzing genomic alterations of the human Rac1 gene. [Copyright &y& Elsevier]

Periodontitis is a chronic inflammatory disease caused by complex interactions between the host immune system and pathogens that affect the integrity of periodontium. To prevent disease progression and thus preserve alveolar bone structure, simultaneous anti-inflammatory and osteogenic intervention are essential. Hence, a glycogen synthase kinase 3 beta inhibitor (BIO) was selected as a potent inflammation modulator and osteogenic agent to achieve this treatment objective. BIO's lack of osteotropicity, poor water solubility, and potential long-term systemic side effects, however, have hampered its clinical applications. To address these limitations, pyrophosphorylated Pluronic F127 (F127-PPi) was synthesized and mixed with regular F127 to prepare an injectable and thermoresponsive hydrogel formulation (PF127) of BIO, which could adhere to hard tissue and gradually release BIO to exert its therapeutic effects locally. Comparing to F127 hydrogel, PF127 hydrogels exhibited stronger binding to hydroxyapatite (HA). Additionally, BIO's solubility in PF127 solution was dramatically improved over F127 solution and the improvement was proportional to the polymer concentration. When evaluated on a rat model of periodontitis, PF127-BIO hydrogel treatment was found to be very effective in preserving alveolar bone and ligament, and preventing periodontal inflammation, as shown by the micro-CT and histological data, respectively. Altogether, these findings suggested that the thermoresponsive PF127 hydrogel is an effective local drug delivery system for better clinical management of periodontitis and associated pathologies. Image 1 [ABSTRACT FROM AUTHOR]