The Role of 11β-Hydroxysteroid Dehydrogenase Type 1 in Wound Healing in Uraemia.

PhD ThesisPeople with Chronic Kidney Disease (CKD), advanced age and systemic cortisol excess demonstrate skin specific similarities including poor wound healing. There is accumulating evidence that patients with advanced kidney disease age prematurely. Studies have demonstrated the cortisol producing 11b-Hydroxysteroid Dehydrogenase Type 1 (11b-HSD-1) enzyme activity is increased in aged skin. We hypothesized the same should be true in the uraemic state. Skin specific 11b-HSD-1 and therefore cortisol excess may be the unifying cause for the skin phenotype and poor healing seen in CKD, advanced age and systemic cortisol excess. The aims of this thesis were to investigate the presence of 11b-HSD-1 and 11b-HSD- 2 within rodent skin with and without uraemia as well as determining the activity of 11b-HSD-1 and to elucidate the role it may play in wound healing. Finally, the efficacy of an 11b-HSD-1 inhibitor in healing was assessed as a potential future therapeutic target. In vitro studies which included Lactate Dehydrogenase (LDH), cell viability, scratch assays and cortisol Enzyme Linked Immunosorbent Assay (ELISA) were conducted using 2 different types of primary skin cultures with the uraemic toxins Indoxyl Sulphate (IS) or p-Cresol (PC) or the specific 11b-HSD-1 inhibitor emodin. An in vivo model of skin wounding was developed in male Wistar rats rendered uraemic by an adenine supplemented diet. Healing was measured in uraemic and non-uraemic control rats at baseline, with the application of topical emodin or with systemic emodin administration. Skin samples were processed ex vivo to extract protein for use in immunoblotting or Ribonucleic Acid (RNA) for Polymerase Chain Reaction (PCR) and a wound healing PCR array. Immunohistochemistry was completed for selected skin samples as were corticosterone (cortisol equivalent in rats) ELISA. The presence of 11b-HSD-1 and 11b-HSD-2 in Human Dermal Fibroblasts (HDF) and Human Epidermal Keratinocytes (HEK) was confirmed by western blotting. No differences in LDH activity, cell viability or cortisol production with the administration of IS, PC or emodin could be detected. Scratch assays utilising uraemic 6 toxins in isolation did not demonstrate a significant delay in wound healing. An improvement in healing using emodin in isolation could also not be demonstrated. Contrary to the in vitro exposure to solitary uraemic toxins, the in vivo findings consistently demonstrated a delay in wound healing in uraemia and the beneficial effect of emodin on healing when administered systemically. This could not be attributed to changes in 11b-HSD-1 or 11b-HSD-2 Messenger RNA (mRNA), protein expression or enzyme activity with the techniques employed. To elucidate a definitive role of 11b-HSD-1 would require the use of radioactive isotopic activity assay and the use of a skin specific 11b-HSD-1 Knock-Out (KO) mouse model. It would be most useful to reproduce this work in human skin, comparing uraemic patients to healthy live kidney donors

Primary vs. Secondary Antibody Deficiency: Clinical Features and Infection Outcomes of Immunoglobulin Replacement

<div><p>Secondary antibody deficiency can occur as a result of haematological malignancies or certain medications, but not much is known about the clinical and immunological features of this group of patients as a whole. Here we describe a cohort of 167 patients with primary or secondary antibody deficiencies on immunoglobulin (Ig)-replacement treatment. The demographics, causes of immunodeficiency, diagnostic delay, clinical and laboratory features, and infection frequency were analysed retrospectively. Chemotherapy for B cell lymphoma and the use of Rituximab, corticosteroids or immunosuppressive medications were the most common causes of secondary antibody deficiency in this cohort. There was no difference in diagnostic delay or bronchiectasis between primary and secondary antibody deficiency patients, and both groups experienced disorders associated with immune dysregulation. Secondary antibody deficiency patients had similar baseline levels of serum IgG, but higher IgM and IgA, and a higher frequency of switched memory B cells than primary antibody deficiency patients. Serious and non-serious infections before and after Ig-replacement were also compared in both groups. Although secondary antibody deficiency patients had more serious infections before initiation of Ig-replacement, treatment resulted in a significant reduction of serious and non-serious infections in both primary and secondary antibody deficiency patients. Patients with secondary antibody deficiency experience similar delays in diagnosis as primary antibody deficiency patients and can also benefit from immunoglobulin-replacement treatment.</p></div

The Role of Toll-like Receptors in Immune Responses Mediated by Human Dendritic Cell Subsets

EThOS - Electronic Theses Online ServiceGBUnited Kingdo

ASPP1 and ASPP2: Common Activators of p53 Family Members

We recently showed that ASPP1 and ASPP2 stimulate the apoptotic function of p53. We show here that ASPP1 and ASPP2 also induce apoptosis independently of p53. By binding to p63 and p73 in vitro and in vivo, ASPP1 and ASPP2 stimulate the transactivation function of p63 and p73 on the promoters of Bax, PIG3, and PUMA but not mdm2 or p21(WAF-1/CIP1). The expression of ASPP1 and ASPP2 also enhances the apoptotic function of p63 and p73 by selectively inducing the expression of endogenous p53 target genes, such as PIG3 and PUMA, but not mdm2 or p21(WAF-1/CIP1). Removal of endogenous p63 or p73 with RNA interference demonstrated that (16) the p53-independent apoptotic function of ASPP1 and ASPP2 is mediated mainly by p63 and p73. Hence, ASPP1 and ASPP2 are the first two identified common activators of all p53 family members. All these results suggest that ASPP1 and ASPP2 could suppress tumor growth even in tumors expressing mutant p53

Diagnostic delay and the presence of bronchiectasis.

<p>Diagnostic delay (time between symptom onset and antibody deficiency diagnosis) was determined for the primary (n = 58) and secondary (n = 25) groups (A). The percentage of subjects with or without bronchiectasis (determined by high-resolution CT scan) is shown for each group (B). Diagnostic delay by bronchiectasis presence or absence is shown for the primary (n = 45) and secondary (n = 21) groups (C). The bars in panels A and C represent median values. Data in panel A were analysed by a two-tailed unequal variance t-test and data in panel C were analysed by a two-tailed Mann-Whitney test; n.s. non-significant (p values <0.05 were considered significant).</p

Immunodeficiency cohort on Ig-replacement treatment.

<p>CVID indicates common variable immune deficiency; ALPS, autoimmune lymphoproliferative syndrome; and WHIM, warts hypogammaglobulinaemia infections and myelokathexis syndrome.</p

Likely cause of secondary antibody deficiency in each subject.

<p>CLL indicates chronic lymphocytic leukaemia; MM, multiple myeloma; MGUS, monoclonal gammopathy of unknown significance; RTX, Rituximab; RA, rheumatoid arthritis; and SLE, systemic lupus erythematosus.</p