Hyaluronan synthases and hyaluronidases in the kidney during changes in hydration status

Hyaluronan is a large glycosaminoglycan that is abundant in the interstitium of the renal medulla/papilla. Papillary hyaluronan increases during hydration and decreases during dehydration. Due to its gel properties and ability to retain large volumes of water, hyaluronan plays a role in renal water handling by affecting the permeability characteristics of the papillary interstitium. The focus of the present investigation was the regulation of hyaluronan metabolism in the kidney, especially during variations in hydration status.In control papillas, HAS 2 mRNA was heavily expressed and HAS 1 and 3 mRNA were weakly distributed. HYALs 1–3 mRNA were found at high expression and HYAL 4 was only weakly expressed. In hydrated animals, the diuretic response (12-fold) was followed by a 58% elevation in papillary hyaluronan and a 45% reduction in the excreted urinary hyaluronidase activity. No difference was determined in HAS 1–3 mRNA or HYAL 1, 3–4 mRNA expression, suggesting a change in activity rather than amount of protein. In dehydrated animals, antidiuresis was followed by a 22% reduction in papillary hyaluronan and a 62% elevation in excreted urinary hyaluronidase activity. Plasma vasopressin was 2.8-fold higher in dehydrated vs. hydrated rats.In conclusion, HAS 2 appears a major contributor to the baseline levels of hyaluronan. Reduced HAS 2 gene expression and increased excreted urinary hyaluronidase activity during dehydration contribute to the reduced amount of hyaluronan and to antidiuretic response.     

Null mutations in Sinorhizobium meliloti exoS and chvI demonstrate the importance of this two‐component regulatory system for symbiosis

Exopolysaccharides, either succinoglycan or galactoglucan, are essential for the establishment of the symbiosis between Sinorhizobium meliloti and Medicago sativa (alfalfa). The ExoS/ChvI two‐component regulatory system is known as a regulator of succinoglycan production but the genes that are directly regulated by ChvI have not been determined. Difficulty isolating exoS and chvI null mutants has prompted the suggestion that these genes are essential for S. meliloti viability. We have successfully isolated exoS and chvI null mutants using a merodiploid‐facilitated strategy. We present evidence that the S. meliloti ExoS/ChvI two‐component regulatory system is essential for symbiosis with alfalfa. Phenotypic analyses of exoS and chvI null mutant strains demonstrate that ExoS/ChvI controls both succinoglycan and galactoglucan production and is required for growth on over 21 different carbon sources. These new findings suggest that the ExoS/ChvI regulatory targets might not be the exo genes that are specific for succinoglycan biosynthesis but rather genes that have common influence on both succinoglycan and galactoglucan production. Other studied alpha‐proteobacteria ExoS/ChvI orthologues are required for the bacteria to invade or persist in host cells and thus we present more evidence that this two‐component regulatory system is essential for alpha‐proteobacterial host interaction.     

Identification of a novel agonist of peroxisome proliferator-activated receptors alpha and gamma that may contribute to the anti-diabetic activity of guggulipid in Lep(ob)/Lep(ob) mice

The ethyl acetate extract of the gum of the guggul tree, Commiphora mukul (guggulipid), is marketed for the treatment of dyslipidaemia and obesity. We have found that it protects Lep(ob)/Lep(ob) mice from diabetes and have investigated possible molecular mechanisms for its metabolic effects, in particular those due to a newly identified component, commipheric acid. Both guggulipid (EC(50)=0.82 microg/ml) and commipheric acid (EC(50)=0.26 microg/ml) activated human peroxisome proliferator-activated receptor alpha (PPARalpha) in COS-7 cells transiently transfected with the receptor and a reporter gene construct. Similarly, both guggulipid (EC(50)=2.3 microg/ml) and commipheric acid (EC(50)=0.3 microg/ml) activated PPARgamma and both promoted the differentiation of 3T3 L1 preadipocytes to adipocytes. Guggulipid (EC(50)=0.66 microg/ml), but not commipheric acid, activated liver X receptor alpha (LXRalpha). E- and Z-guggulsterones, which are largely responsible for guggulipid's hypocholesterolaemic effect, had no effects in these assays. Guggulipid (20 g/kg diet) improved glucose tolerance in female Lep(ob)/Lep(ob) mice. Pure commipheric acid, given orally (960 mg/kg body weight, once daily), increased liver weight but did not affect body weight or glucose tolerance. However, the ethyl ester of commipheric acid (150 mg/kg, twice daily) lowered fasting blood glucose and plasma insulin, and plasma triglycerides without affecting food intake or body weight. These results raise the possibility that guggulipid has anti-diabetic activity due partly to commipheric acid's PPARalpha/gamma agonism, but the systemic bioavailability of orally dosed, pure commipheric acid appears poor. Another component may contribute to guggulipid's anti-diabetic and hypocholesterolaemic activity by stimulating LXRalpha.     

Sonic hedgehog signalling inhibits palatogenesis and arrests tooth development in a mouse model of the nevoid basal cell carcinoma syndrome

Nevoid basal cell carcinoma syndrome (NBCCS) is an autosomal dominant or spontaneous disorder characterized by multiple cutaneous basal cell carcinomas, odontogenic keratocysts, skeletal anomalies and facial dysmorphology, including cleft lip and palate. Causative mutations for NBCCS occur in the PTCH1 gene on chromosome 9q22.3-q31, which encodes the principle receptor for the Hedgehog signalling pathway. We have investigated the molecular basis of craniofacial defects seen in NBCCS using a transgenic mouse model expressing Shh in basal epithelium under a Keratin-14 promoter. These mice have an absence of flat bones within the skull vault, hypertelorism, open-bite malocclusion, cleft palate and arrested tooth development. Significantly, increased Hedgehog signal transduction in these mice can influence cell fate within the craniofacial region. In medial edge epithelium of the palate, Shh activity prevents apoptosis and subsequent palatal shelf fusion. In contrast, high levels of Shh in odontogenic epithelium arrests tooth development at the bud stage, secondary to a lack of cell proliferation in this region. These findings illustrate the importance of appropriately regulated Hedgehog signalling during early craniofacial development and demonstrate that oro-facial clefting and hypodontia seen in NBCCS can occur as a direct consequence of increased Shh signal activity within embryonic epithelial tissues.     

The role of amphibian antimicrobial peptides in protection of amphibians from pathogens linked to global amphibian declines

Amphibian species have experienced population declines and extinctions worldwide that are unprecedented in recent history. Many of these recent declines have been linked to a pathogenic skin fungus, Batrachochytrium dendrobatidis, or to iridoviruses of the genus Ranavirus. One of the first lines of defense against pathogens that enter by way of the skin are antimicrobial peptides synthesized and stored in dermal granular glands and secreted into the mucus following alarm or injury. Here, I review what is known about the capacity of amphibian antimicrobial peptides from diverse amphibians to inhibit B. dendrobatidis or ranavirus infections. When multiple species were compared for the effectiveness of their in vitro antimicrobial peptides defenses against B. dendrobatidis, non-declining species of rainforest amphibians had more effective antimicrobial peptides than species in the same habitat that had recently experienced population declines. Further, there was a significant correlation between the effectiveness of the antimicrobial peptides and resistance of the species to experimental infection. These studies support the hypothesis that antimicrobial peptides are an important component of innate defenses against B. dendrobatidis. Some amphibian antimicrobial peptides inhibit ranavirus infections and infection of human T lymphocytes by the human immunodeficiency virus (HIV). An effective antimicrobial peptide defense against skin pathogens appears to depend on a diverse array of genes expressing antimicrobial peptides. The production of antimicrobial peptides may be regulated by signals from the pathogens. However, this defense must also accommodate potentially beneficial microbes on the skin that compete or inhibit growth of the pathogens. How this delicate balancing act is accomplished is an important area of future research.     

Identification of a mutation in the para-sodium channel gene of the cattle tick Rhipicephalus (Boophilus) microplus associated with resistance to synthetic pyrethroid acaricides

Resistance against synthetic pyrethroid (SP) products for the control of cattle ticks in Australia was detected in the field in 1984, within a very short time of commercial introduction. We have identified a mutation in the domain II S4-5 linker of the para-sodium channel that is associated with resistance to SPs in the cattle tick Rhipicephalus (Boophilus) microplus from Australia. The cytosine to adenine mutation at position 190 in the R. microplus sequence AF134216, results in an amino acid substitution from leucine in the susceptible strain to isoleucine in the resistant strain. A similar mutation has been shown to confer SP resistance in the whitefly, Bemisia tabaci, but has not been described previously in ticks. A diagnostic quantitative PCR assay has been developed using allele-specific Taqman^® minor groove-binding (MGB) probes. Using the assay to screen field and laboratory populations of ticks showed that homozygote allelic frequencies correlated highly with the survival percentage at the discriminating concentration of cypermethrin.     

Effect of conformity and contact stress on wear in fixed-bearing total knee prostheses

Ultra high molecular weight polyethylene (PE) remains the primary bearing surface of choice in total knee replacements (TKR). Wear is controlled by levels of cross-shear motion and contact stress. The aim of this study was to compare the wear of fixed-bearing total knee replacements with curved and flat inserts and to test the hypothesis that the flat inserts which give higher contact stresses and smaller contact areas would lead to lower levels of surface wear. A low-conforming, high contact stress knee with a low-medium level of cross shear resulted in significantly lower wear rates in comparison to a standard cruciate sacrificing fixed-bearing knee. The low wear solution found in the knee simulator was supported by fundamental studies of wear as a function of pressure and cross shear in the pin on plate system. Current designs of fixed-bearing knees do not offer this low wear solution due to their medium cross shear, moderate conformity and medium contact stress.     

Secreted Protein Acidic and Rich in Cysteine (SPARC) in Human Skeletal Muscle

Secreted protein acidic and rich in cysteine (SPARC)/osteonectin is expressed in different tissues during remodeling and repair, suggesting a function in regeneration. Several gene expression studies indicated that SPARC was expressed in response to muscle damage. Studies on myoblasts further indicated a function of SPARC in skeletal muscle. We therefore found it of interest to study SPARC expression in human skeletal muscle during development and in biopsies from Duchenne and Becker muscular dystrophy and congenital muscular dystrophy, congenital myopathy, inclusion body myositis, and polymyositis patients to analyze SPARC expression in a selected range of inherited and idiopathic muscle wasting diseases. SPARC-positive cells were observed both in fetal and neonatal muscle, and in addition, fetal myofibers were observed to express SPARC at the age of 15–16 weeks. SPARC protein was detected in the majority of analyzed muscle biopsies (23 of 24), mainly in mononuclear cells of which few were pax7 positive. Myotubes and regenerating myofibers also expressed SPARC. The expression-degree seemed to reflect the severity of the lesion. In accordance with these in vivo findings, primary human-derived satellite cells were found to express SPARC both during proliferation and differentiation in vitro. In conclusion, this study shows SPARC expression both during muscle development and in regenerating muscle. The expression is detected both in satellite cells/myoblasts and in myotubes and muscle fibers, indicating a role for SPARC in the skeletal muscle compartment. (J Histochem Cytochem 57:29–39, 2009)