Vascularized acellular dermal matrix island flaps for the repair of abdominal muscle defects

The potential widespread use of tissue-engineered matrices in soft-tissue reconstruction has been limited by the difficulty in fabricating and confirming a functional microcirculation. Acellular dermal matrix placed in a soft-tissue pocket acts as a scaffold to be incorporated by the host's fibrovascular tissue. A new method for noninvasive real-time observation of functional microvascular networks using orthogonal polarization spectral (OPS) imaging has recently been reported. Arterioles, venules, and capillaries can be directly visualized, and the movement of individual blood cells through them can be observed. The present study was performed to investigate the use of prefabricated acellular dermal matrix with an arteriovenous unit for the repair of abdominal muscle defects. OPS imaging was used to determine the presence of a functional microcirculation in the neovascularized matrix. In Sprague-Dawley rats, vascularized matrix was prefabricated by placing the superficial epigastric artery and vein on a 2-cm x 2-cm implant-type acellular dermal matrix in the thigh. Three weeks after implantation, the matrix-arteriovenous unit was elevated as an axial-type flap and a 2-cm x 2-cm full-thickness block of abdominal muscle immediately superior to the inguinal ligament was resected. Additional procedures were performed according to group: no repair (group 1, n = 20); repair with nonvascularized acellular dermal matrix (group 2, n = 20); repair with devascularized acellular dermal matrix (group 3, = 20); and repair with vascularized acellular dermal matrix (group 4, n = 20). OPS imaging (field of view, 1 mm in diameter; scan depth range, 0.2 mm) was performed on both sides of each flap on a total of 10 random distal regions before and after pedicle transection in group 3 and with the pedicle preserved in group 4. Hernia rate and duration of survival were compared for 21 days. OPS imaging showed directional blood cell movement through the capillary network in all areas scanned in group 4. No microvascular perfusion was observed after pedicle transection in group 3. Hernia rates of 100, 80, 90, and 0 percent were seen in groups 1, 2, 3, and 4, respectively. Median survival times of 9, 11.5, 9, and 21 postoperative days were noted in groups 1, 2, 3, and 4, respectively. Histopathologic analysis with factor VIII revealed full-thickness infiltration of the matrix by endothelial cells, signifying newly formed blood vessels. Repair of abdominal muscle defects using vascularized acellular dermal matrix resulted in no hernia and survival of all animals for the duration of study. However, repairs using avascular or devascularized matrix resulted in significant rates of hernia and decreased survival. Acellular dermal matrix can be prefabricated into vascularized tissue using an arteriovenous unit and used successfully to repair abdominal muscle defects. OPS imaging allowed for high-contrast direct visualization of microcirculation in previously acellular tissue following prefabrication with an arteriovenous unit.     

Targeting tuberculosis and malaria through inhibition of Enoyl reductase: compound activity and structural data

Tuberculosis and malaria together result in an estimated 5 million deaths annually. The spread of multidrug resistance in the most pathogenic causative agents, Mycobacterium tuberculosis and Plasmodium falciparum, underscores the need to identify active compounds with novel inhibitory properties. Although genetically unrelated, both organisms use a type II fatty-acid synthase system. Enoyl acyl carrier protein reductase (ENR), a key type II enzyme, has been repeatedly validated as an effective antimicrobial target. Using high throughput inhibitor screens with a combinatorial library, we have identified two novel classes of compounds with activity against the M. tuberculosis and P. falciparum enzyme (referred to as InhA and PfENR, respectively). The crystal structure of InhA complexed with NAD+ and one of the inhibitors was determined to elucidate the mode of binding. Structural analysis of InhA with the broad spectrum antimicrobial triclosan revealed a unique stoichiometry where the enzyme contained either a single triclosan molecule, in a configuration typical of other bacterial ENR:triclosan structures, or harbored two triclosan molecules bound to the active site. Significantly, these compounds do not require activation and are effective against wild-type and drug-resistant strains of M. tuberculosis and P. falciparum. Moreover, they provide broader chemical diversity and elucidate key elements of inhibitor binding to InhA for subsequent chemical optimization.     

Opt-out of voluntary HIV testing: a Singapore hospital's experience

Introduction

Since 2008, the Singapore Ministry of Health (MOH) has expanded HIV testing by increasing anonymous HIV test sites, as well as issuing a directive to hospitals to offer routine voluntary opt out inpatient HIV testing. We reviewed this program implemented at the end of 2008 at Tan Tock Seng Hospital (TTSH), the second largest acute care general hospital in Singapore.

Methods and Findings

From January 2009 to December 2010, all inpatients aged greater or equal than 21 years were screened for HIV unless they declined or were not eligible for screening. We reviewed the implementation of the Opt Out testing policy. There were a total of 93,211 admissions; 41,543 patients were included based on HIV screening program eligibility criteria. Among those included, 79% (n = 32,675) opted out of HIV screening. The overall acceptance rate was 21%. Majority of eligible patients who were tested (63%) were men. The mean age of tested patients was 52 years. The opt out rate was significantly higher among females (OR: 1.5, 95%CI: 1.4–1.6), aged >60 years (OR: 2.3, 95%CI: 2.2–2.4) and Chinese ethnicity (OR: 1.7, 95%CI:1.6–1.8). The false positive rate of the HIV screening test is 0.56%. The proportion of patients with HIV infection among those who underwent HIV screening is 0.18%. All16 confirmed HIV patients were linked to care.

Conclusion

The default opt-in rate of inpatient HIV testing was low at Tan Tock Seng Hospital, Singapore. Efforts to address individual HIV risk perception and campaigns against HIV stigma are needed to encourage more individuals to be tested for HIV.     

Intestinal redox biology and oxidative stress

The intestinal epithelium sits at the interface between an organism and its luminal environment, and as such is prone to oxidative damage induced by luminal oxidants. Mucosal integrity is maintained by the luminal redox status of the glutathione/glutathione disulfide (GSH/GSSG) and cysteine/cystine (Cys/CySS) couples which also support luminal nutrient absorption, mucus fluidity, and a diverse microbiota. The epithelial layer is uniquely organized for rapid self-renewal that is achieved by the well-regulated processes of crypt stem cell proliferation and crypt-to-villus cell differentiation. The GSH/GSSG and Cys/CySS redox couples, known to modulate intestinal cell transition through proliferation, differentiation or apoptosis, could govern the regenerative potential of the mucosa. These two couples, together with that of the thioredoxin/thioredoxin disulfide (Trx/TrxSS) couple are the major intracellular redox systems, and it is proposed that they each function as distinctive redox control nodes or circuitry in the control of metabolic processes and networks of enzymatic reactions. Specificity of redox signaling is accomplished in part by subcellular compartmentation of the individual redox systems within the mitochondria, nucleus, endoplasmic reticulum, and cytosol wherein each defined redox environment is suited to the specific metabolic function within that compartment. Mucosal oxidative stress would result from the disruption of these unique redox control nodes, and the subsequent alteration in redox signaling can contribute to the development of degenerative pathologies of the intestine, such as inflammation and cancer.     

Interaction of the heart-specific LIM domain protein, FHL2, with DNA-binding nuclear protein, hNP220

Using a yeast two-hybrid library screen, we have identified that the heart specific FHL2 protein, four-and-a-half LIM protein 2, interacted with human DNA-binding nuclear protein, hNP220. Domain studies by the yeast two-hybrid interaction assay revealed that the second LIM domain together with the third and the fourth LIM domains of FHL2 were responsible to the binding with hNP220. Using green fluorescent protein (GFP)-FHL2 and blue fluorescent protein (BFP)-hNP220 fusion proteins co-expressed in the same cell, we demonstrated a direct interaction between FHL2 and hNP220 in individual nucleus by two-fusion Fluorescence Resonance Energy Transfer (FRET) assay. Besides, Western blot analysis using affinity-purified anti-FHL2 antipeptide antibodies confirmed a 32-kDa protein of FHL2 in heart only. Virtually no expression of FHL2 protein was detected in brain, liver, lung, kidney, testis, skeletal muscle, and spleen. Moreover, the expression of FHL2 protein was also detectable in the human diseased heart tissues. Our results imply that FHL2 protein can shuttle between cytoplasm and nucleus and may act as a molecular adapter to form a multicomplex with hNP220 in the nucleus, thus we speculate that FHL2 may be particularly important for heart muscle differentiation and the maintenance of the heart phenotype.     

Role of the histone H3 lysine 4 methyltransferase, SET7/9, in the regulation of NF-kappaB-dependent inflammatory genes. Relevance to diabetes and inflammation

Nuclear factor kappa-B (NF-kappaB)-regulated inflammatory genes, such as TNF-alpha (tumor necrosis factor-alpha), play key roles in the pathogenesis of inflammatory diseases, including diabetes and the metabolic syndrome. However, the nuclear chromatin mechanisms are unclear. We report here that the chromatin histone H3-lysine 4 methyltransferase, SET7/9, is a novel coactivator of NF-kappaB. Gene silencing of SET7/9 with small interfering RNAs in monocytes significantly inhibited TNF-alpha-induced inflammatory genes and histone H3-lysine 4 methylation on these promoters, as well as monocyte adhesion to endothelial or smooth muscle cells. Chromatin immunoprecipitation revealed that SET7/9 small interfering RNA could reduce TNF-alpha-induced recruitment of NF-kappaB p65 to inflammatory gene promoters. Inflammatory gene induction by ligands of the receptor for advanced glycation end products was also attenuated in SET7/9 knockdown monocytes. In addition, we also observed increased inflammatory gene expression and SET7/9 recruitment in macrophages from diabetic mice. Microarray profiling revealed that, in TNF-alpha-stimulated monocytes, the induction of 25% NF-kappaB downstream genes, including the histone H3-lysine 27 demethylase JMJD3, was attenuated by SET7/9 depletion. These results demonstrate a novel role for SET7/9 in inflammation and diabetes.     

Transformation of galena to pyromorphite produces bioavailable sulfur for neutrophilic chemoautotrophy

The aqueous concentration of lead [Pb(II)] in geochemical environments is controlled by the solubility of Pb‐bearing minerals and their weathering products. In contaminated soils, a common method for in situ stabilization of Pb(II) is the addition of phosphate to convert more redox sensitive sulfide minerals into sparingly soluble pyromorphite [Pb₅(PO₄)₃X]. In this study, we conducted experimental studies to investigate the fate of reduced sulfur during the conversion of galena [PbS] to chloropyromorphite [Pb₅(PO₄)₃Cl]. Powder X‐ray diffraction analysis indicated that the reaction of phosphate with galena under oxic conditions resulted in the oxidation of sulfide and formation of elemental sulfur [S₈]. Under oxic abiotic conditions, the S₈ was retained in the solid phase, and negligible concentrations of sulfur as sulfide and thiosulfate were detected in the aqueous phase and only a small amount of sulfate. When PbS reacted in the presence of the chemoautotrophic organism Bosea sp. WAO, the S₈ in the secondary mineral was oxidized to sulfate. Strain WAO produced significantly more sulfate from the secondary S₈ than from the primary galena. Microscopic analysis of mineral–microbe aggregates on mineral‐embedded slide cultures showed that the organism was colocalized and increased in biomass over time on the secondary mineral surface supporting a microbial role. The results of this study indicate that stimulation of sulfur‐oxidizing activity may be a direct consequence of phosphate amendments to Pb(II)‐contaminated soils.     

Haptic guidance improves the visuo-manual tracking of trajectories

BACKGROUND: Learning to perform new movements is usually achieved by following visual demonstrations. Haptic guidance by a force feedback device is a recent and original technology which provides additional proprioceptive cues during visuo-motor learning tasks. The effects of two types of haptic guidances-control in position (HGP) or in force (HGF)-on visuo-manual tracking ("following") of trajectories are still under debate. METHODOLOGY/PRINCIPALS FINDINGS: Three training techniques of haptic guidance (HGP, HGF or control condition, NHG, without haptic guidance) were evaluated in two experiments. Movements produced by adults were assessed in terms of shapes (dynamic time warping) and kinematics criteria (number of velocity peaks and mean velocity) before and after the training sessions. Trajectories consisted of two Arabic and two Japanese-inspired letters in Experiment 1 and ellipses in Experiment 2. We observed that the use of HGF globally improves the fluency of the visuo-manual tracking of trajectories while no significant improvement was found for HGP or NHG. CONCLUSION/SIGNIFICANCE: These results show that the addition of haptic information, probably encoded in force coordinates, play a crucial role on the visuo-manual tracking of new trajectories.