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Acute graft-versus-host disease (aGVHD) and kidney injury are the major complications after allogeneic hematopoietic stem cell transplantation (HSCT). Although the underlying mechanisms for the development of these complications are not yet fully understood, it has been proposed that emergence of aGVHD contributes to the development of kidney injury after HSCT. We have shown previously that aGVHD targets the kidney in a biphasic manner: at the onset, inflammatory genes are up-regulated, while when aGVHD becomes established, donor lymphocytes infiltrate the kidney. Here, we characterize renal manifestations at the onset of aGVHD. Mice receiving allogeneic bone marrow and spleen cells displayed symptoms of aGVHD and elevated serum levels of tumor necrosis factor alpha (TNF-α) and interferon gamma (IFN-γ) within 4 days. There was concurrent kidney injury with the following characteristics: (1) elevated expression of the kidney injury biomarker, neutrophil gelatinase-associated lipocalin (NGAL), (2) accumulation of hetero-lysosomes in proximal tubule epithelial cells, and (3) reductions in αKlotho mRNA and protein and increased serum levels of fibroblast growth factor 23 (Fgf23), phosphate and urea. This situation resembled acute renal injury caused by bacterial lipopolysaccharide. We conclude that the onset of aGVHD is associated with kidney injury involving down-regulation of αKlotho, a sight that may inspire novel therapeutic approaches.

Calcium and phosphorous are critical elements in a number of physiological processes, including maintenance of bone structure, cell signaling and energy metabolism. Their endocrine regulation is tightly controlled through a number of feedback mechanisms involving parathyroid hormone (PTH), vitamin D and the recently discovered fibroblast growth factor 23 (FGF23). The kidney is a key organ in maintaining normal serum levels of calcium and inorganic phosphorous, and disturbances in mineral metabolism are commonly observed in patients with chronic kidney disease (CKD). Klotho is a membrane-bound protein expressed in the renal tubules that acts as a co-receptor for FGF23. In addition, Klotho can be shedded from the cell surface to extra-cellular compartments and function as a hormone with effects on mineral metabolism independent of FGF23. During the progression of CKD the expression of Klotho rapidly declines, and accumulating evidence point to lack of Klotho as a pathogenic factor driving clinical complications in CKD. The main focus of this thesis has been to elucidate the role of renal Klotho in mineral metabolism and on systemic effects. In Study I we generated distal tubule-specific Klotho knockout mice (Ksp-KL-/-) by employing cre-lox recombination. Ksp-KL-/- mice were hyperphosphatemic with elevated serum Fgf23 levels, indicating that distal tubular Klotho affects phosphate reabsorption in the proximal tubules. The exact mechanism of this proposed distal-toproximal tubular signaling remains unknown. In Study II we generated mice with Klotho deleted throughout the nephron (Six2-KL-/- ). Six2-KL-/- mice were infertile, kyphotic, growth retarded and had a decreased life span, closely resembling the phenotype seen in systemic Klotho knockout mice. Also the serum and urine biochemistries, low serum Klotho levels as well as profound histological abnormalities were indistinguishable from systemic Klotho knockout mice, unraveling the kidney as the principle contributor

BACKGROUND: Epidemiological evidence indicates yet unknown epigenetic mechanisms underlying a propensity for overweight and type 2 diabetes. We analyzed the extent of methylation at lysine 4 and lysine 9 of histone H3 in primary human adipocytes from 43 subjects using modification-specific antibodies. RESULTS: The level of lysine 9 dimethylation was stable, while adipocytes from type 2 diabetic and non-diabetic overweight subjects exhibited about 40% lower levels of lysine 4 dimethylation compared with cells from normal-weight subjects. In contrast, trimethylation at lysine 4 was 40% higher in adipocytes from overweight diabetic subjects compared with normal-weight and overweight non-diabetic subjects. There was no association between level of modification and age of subjects. CONCLUSIONS: The findings define genome-wide molecular modifications of histones in adipocytes that are directly associated with overweight and diabetes, and thus suggest a molecular basis for existing epidemiological evidence of epigenetic inheritance.

INTRODUCTION: mTOR and its downstream effectors the 4E-binding protein 1 (4EBP1) and the p70 ribosomal S6 kinases (S6K1 and S6K2) are frequently upregulated in breast cancer, and assumed to be driving forces in tumourigenesis, in close connection with oestrogen receptor (ER) networks. Here, we investigated these factors as clinical markers in five different cohorts of breast cancer patients. METHODS: The prognostic significance of 4EBP1, S6K1 and S6K2 mRNA expression was assessed with real-time PCR in 93 tumours from the treatment randomised Stockholm trials, encompassing postmenopausal patients enrolled between 1976 and 1990. Three publicly available breast cancer cohorts were used to confirm the results. Furthermore, the predictive values of 4EBP1 and p4EBP1_S65 protein expression for both prognosis and endocrine treatment benefit were assessed by immunohistochemical analysis of 912 node-negative breast cancers from the Stockholm trials. RESULTS: S6K2 and 4EBP1 mRNA expression levels showed significant correlation and were associated with a poor outcome in all cohorts investigated. 4EBP1 protein was confirmed as an independent prognostic factor, especially in progesterone receptor (PgR)-expressing cancers. 4EBP1 protein expression was also associated with a poor response to endocrine treatment in the ER/PgR positive group. Cross-talk to genomic as well as non-genomic ER/PgR signalling may be involved and the results further support a combination of ER and mTOR signalling targeted therapies. CONCLUSION: This study suggests S6K2 and 4EBP1 as important factors for breast tumourigenesis, interplaying with hormone receptor signalling. We propose S6K2 and 4EBP1 as new potential clinical markers for prognosis and endocrine therapy response in breast cancer.