Emma Tham

Antimicrobial protein hCAP18/LL-37 is highly expressed in breast cancer and is a putative growth factor for epithelial cells

Human cathelicidin antimicrobial protein hCAP18/LL‐37 is an effector molecule of the nonspecific innate immune system. hCAP18/LL‐37 is present in leukocytes and is expressed in skin and other epithelia, where it is upregulated in association with inflammation and injury. In addition, antimicrobial proteins including cathelicidins have been proposed to play a role in the nonspecific defense against tumors. To assess its potential role in tumor host defense, we investigated the expression of hCAP18/LL‐37 in a series of breast carcinomas. Unexpectedly, we found that hCAP18/LL‐37 was strongly expressed in the tumor cells and not in the adjacent stroma. To test the hypothesis that hCAP18/LL‐37 may provide a growth advantage for the tumor cells, we treated human epithelial cell lines with synthetic biologically active LL‐37 peptide and found a significant increase in cell proliferation. In addition, transgenic expression of hCAP18 in 2 different human epithelial cell lines resulted in increased proliferation of both cell types. These findings do not support the hypothesis that LL‐37 has an antitumor effect, but rather suggest that hCAP18/LL‐37 may promote tumor cell growth in breast cancer.

Vascular endothelial growth factor is highly expressed in muscle tissue of patients with polymyositis and patients with dermatomyositis.

OBJECTIVE To investigate the expression of vascular endothelial growth factor (VEGF) in muscle biopsy specimens and serum from patients with polymyositis and patients with dermatomyositis compared with that in healthy control subjects. METHODS Muscle biopsy specimens from 33 patients with polymyositis or dermatomyositis and 15 healthy control subjects and serum samples from 56 patients and 56 healthy control subjects were analyzed. Patients were categorized into 3 groups, depending on disease duration and the presence or absence of inflammatory infiltrates. The expression of VEGF and the vessel marker CD31 in muscle was analyzed by immunohistochemistry, the expression of VEGF messenger RNA (mRNA) was analyzed by in situ hybridization, and serum levels of VEGF were determined by enzyme-linked immunosorbent assay. RESULTS Patients with polymyositis or dermatomyositis in the early or chronic phase without inflammatory infiltrates had a decreased total number of capillaries compared with healthy individuals. In patients with early disease without inflammatory infiltrates, the number of VEGF-expressing muscle fibers was increased compared with that in control subjects, whereas VEGF expression was unchanged in the chronic phase of disease. In patients with established disease with inflammatory infiltrates, total VEGF expression was high compared with that in healthy control subjects. In healthy control subjects, VEGF was expressed in endothelial cells and in occasional muscle fibers. VEGF mRNA was expressed in muscle fibers in both healthy individuals and patients. The level of serum VEGF was significantly increased in patients compared with control subjects. CONCLUSION Our observations support a role of VEGF in the early phases of polymyositis and dermatomyositis. A reduced number of capillaries could lead to induction of VEGF expression in muscle fibers. Furthermore, differences in molecular expression during certain phases of disease may help in the development of specific therapeutic algorithms in the treatment of myositis.

Dominant Mutations in KAT6A Cause Intellectual Disability with Recognizable Syndromic Features

Through a multi-center collaboration study, we here report six individuals from five unrelated families, with mutations in KAT6A/MOZ detected by whole-exome sequencing. All five different de novo heterozygous truncating mutations were located in the C-terminal transactivation domain of KAT6A: NM_001099412.1: c.3116_3117 delCT, p.(Ser1039∗); c.3830_3831insTT, p.(Arg1278Serfs∗17); c.3879 dupA, p.(Glu1294Argfs∗19); c.4108G>T p.(Glu1370∗) and c.4292 dupT, p.(Leu1431Phefs∗8). An additional subject with a 0.23 MB microdeletion including the entire KAT6A reading frame was identified with genome-wide array comparative genomic hybridization. Finally, by detailed clinical characterization we provide evidence that heterozygous mutations in KAT6A cause a distinct intellectual disability syndrome. The common phenotype includes hypotonia, intellectual disability, early feeding and oromotor difficulties, microcephaly and/or craniosynostosis, and cardiac defects in combination with subtle facial features such as bitemporal narrowing, broad nasal tip, thin upper lip, posteriorly rotated or low-set ears, and microretrognathia. The identification of human subjects complements previous work from mice and zebrafish where knockouts of Kat6a/kat6a lead to developmental defects.

Decreased Expression of VEGF-A in Rat Experimental Autoimmune Encephalomyelitis and in Cerebrospinal Fluid Mononuclear Cells from Patients with Multiple Sclerosis

Vascular endothelial growth factor A (VEGF‐A) stimulates angiogenesis, but is also pro‐inflammatory and plays an important role in the development of neurological disease, where it can have both attenuating and exacerbating effects. VEGF‐B, a related molecule, is highly expressed in the central nervous system and seems to be important in neurological injury. A few studies have indicated that VEGF‐A may play a role in the pathogenesis of multiple sclerosis (MS), but the role of VEGF‐B has not been studied. We have studied the expression of VEGF‐A, ‐B and their receptors by mRNA in situ hybridization, immunohistochemistry and real‐time PCR in spinal cord from LEW rats with experimental autoimmune encephalomyelitis (EAE) and in cerebrospinal fluid (CSF) and blood samples from MS patients. Whereas VEGF‐A is downregulated in glia in EAE, the infiltrating inflammatory cells are positive for VEGF‐A. Expression of VEGF‐B and the VEGF receptors is unaltered. In addition, the levels of VEGF‐A mRNA in monocytes in CSF are lower in MS patients compared with controls. These results demonstrate a complex regulation of VEGF‐A during neuroinflammation and suggest that VEGF‐B is not involved in the pathogenesis of MS.

Predicting the risk of multiple endocrine neoplasia type 1 for patients with commonly occurring endocrine tumors

OBJECTIVE Endocrine diseases that can be part of the rare inheritable syndrome multiple endocrine neoplasia type 1 (MEN1) commonly occur in the general population. Patients at risk for MEN1, and consequently their families, must be identified to prevent morbidity through periodic screening for the detection and treatment of manifestations in an early stage. The aim of the study was to develop a model for predicting MEN1 in individual patients with sporadically occurring endocrine tumors. DESIGN Cross-sectional study. METHODS In a nationwide study in The Netherlands, patients with sporadically occurring endocrine tumors in whom the referring physician suspected the MEN1 syndrome were identified between 1998 and 2011 (n=365). Logistic regression analysis with internal validation using bootstrapping and external validation with a cohort from Sweden was used. RESULTS A MEN1 mutation was found in 15.9% of 365 patients. Recurrent primary hyperparathyroidism (pHPT; odds ratio (OR) 162.40); nonrecurrent pHPT (OR 25.78); pancreatic neuroendocrine tumors (pNETs) and duodenal NETs (OR 17.94); pituitary tumor (OR 4.71); NET of stomach, thymus, or bronchus (OR 25.84); positive family history of NET (OR 4.53); and age (OR 0.96) predicted MEN1. The c-statistic of the prediction model was 0.86 (95% confidence interval (95% CI) 0.81-0.90) in the derivation cohort and 0.77 (95% CI 0.66-0.88) in the validation cohort. CONCLUSION With the prediction model, the risk of MEN1 can be calculated in patients suspected for MEN1 with sporadically occurring endocrine tumors.

Upregulation of VEGF-A Without Angiogenesis in a Mouse Model of Dilated Cardiomyopathy Caused by Mitochondrial Dysfunction

Angiogenesis is implicated in a variety of human pathologies and may also play a role in the progression of heart failure. We have studied the expression of members of the vascular endothelial growth factor (VEGF) and the angiopoietin families and their receptors in mice lacking the mitochondrial transcription factor A. These mice lack functional respiratory chain activity in their myocytes and develop dilated cardiomyopathy (DCM) postnatally. We studied the hearts of the knockout mice by in situ hybridization, Western blotting analysis, and immunohistochemistry. VEGF-A mRNA and protein levels were elevated in the myocardium of the knockouts. Levels of the hypoxia inducible transcription factor 1 alpha (HIF1α) and of glyceraldehyde-3-phosphate dehydrogenase transcripts were also increased, whereas those of angiopoietin−1 and −2 were reduced. Despite the striking upregulation of VEGF-A, there was no increase in capillary density in the knockout hearts. This study suggests that a disturbance in angiogenesis may contribute to the pathogenesis of DCM.

Five endometrial cancer risk loci identified through genome-wide association analysis

We conducted a meta-analysis of three endometrial cancer genome-wide association studies (GWAS) and two follow-up phases totaling 7,737 endometrial cancer cases and 37,144 controls of European ancestry. Genome-wide imputation and meta-analysis identified five new risk loci of genome-wide significance at likely regulatory regions on chromosomes 13q22.1 (rs11841589, near KLF5), 6q22.31 (rs13328298, in LOC643623 and near HEY2 and NCOA7), 8q24.21 (rs4733613, telomeric to MYC), 15q15.1 (rs937213, in EIF2AK4, near BMF) and 14q32.33 (rs2498796, in AKT1, near SIVA1). We also found a second independent 8q24.21 signal (rs17232730). Functional studies of the 13q22.1 locus showed that rs9600103 (pairwise r2 = 0.98 with rs11841589) is located in a region of active chromatin that interacts with the KLF5 promoter region. The rs9600103[T] allele that is protective in endometrial cancer suppressed gene expression in vitro, suggesting that regulation of the expression of KLF5, a gene linked to uterine development, is implicated in tumorigenesis. These findings provide enhanced insight into the genetic and biological basis of endometrial cancer.

The Expression of VEGF-A Is Down Regulated in Peripheral Blood Mononuclear Cells of Patients with Secondary Progressive Multiple Sclerosis

Background Most patients with relapsing-remitting multiple sclerosis (RRMS) eventually enter a secondary progressive (SPMS) phase, characterized by increasing neurological disability. The mechanisms underlying transition to SPMS are unknown and effective treatments and biomarkers are lacking. Vascular endothelial growth factor-A (VEGF-A) is an angiogenic factor with neuroprotective effects that has been associated with neurodegenerative diseases. SPMS has a prominent neurodegenerative facet and we investigated a possible role for VEGF-A during transition from RRMS to SPMS. Methodology/Principal Findings VEGF-A mRNA expression in peripheral blood mononuclear (PBMC) and cerebrospinal fluid (CSF) cells from RRMS (n = 128), SPMS (n = 55) and controls (n = 116) were analyzed using real time PCR. We demonstrate reduced expression of VEGF-A mRNA in MS CSF cells compared to controls (p<0.001) irrespective of disease course and expression levels are restored by natalizumab treatment(p<0.001). VEGF-A was primarily expressed in monocytes and our CSF findings in part may be explained by effects on relative monocyte proportions. However, VEGF-A mRNA expression was also down regulated in the peripheral compartment of SPMS (p<0.001), despite unchanged monocyte counts, demonstrating a particular phenotype differentiating SPMS from RRMS and controls. A possible association of allelic variability in the VEGF-A gene to risk of MS was also studied by genotyping for six single nucleotide polymorphisms (SNPs) in MS (n = 1114) and controls (n = 1234), which, however, did not demonstrate any significant association between VEGF-A alleles and risk of MS. Conclusions/Significance Expression of VEGF-A in CSF cells is reduced in MS patients compared to controls irrespective of disease course. In addition, SPMS patients display reduced VEGF-A mRNA expression in PBMC, which distinguish them from RRMS and controls. This indicates a possible role for VEGF-A in the mechanisms regulating transition to SPMS. Decreased levels of PBMC VEGF-A mRNA expression should be further evaluated as a biomarker for SPMS.

CYP19A1 fine-mapping and Mendelian randomization: estradiol is causal for endometrial cancer

Candidate gene studies have reported CYP19A1 variants to be associated with endometrial cancer and with estradiol (E2) concentrations. We analyzed 2937 single nucleotide polymorphisms (SNPs) in 6608 endometrial cancer cases and 37 925 controls and report the first genome wide-significant association between endometrial cancer and a CYP19A1 SNP (rs727479 in intron 2, P=4.8×10−11). SNP rs727479 was also among those most strongly associated with circulating E2 concentrations in 2767 post-menopausal controls (P=7.4×10−8). The observed endometrial cancer odds ratio per rs727479 A-allele (1.15, CI=1.11–1.21) is compatible with that predicted by the observed effect on E2 concentrations (1.09, CI=1.03–1.21), consistent with the hypothesis that endometrial cancer risk is driven by E2. From 28 candidate-causal SNPs, 12 co-located with three putative gene-regulatory elements and their risk alleles associated with higher CYP19A1 expression in bioinformatical analyses. For both phenotypes, the associations with rs727479 were stronger among women with a higher BMI (Pinteraction=0.034 and 0.066 respectively), suggesting a biologically plausible gene-environment interaction.

A novel phenotype in N-glycosylation disorders: Gillessen-Kaesbach-Nishimura skeletal dysplasia due to pathogenic variants in ALG9.

A rare lethal autosomal recessive syndrome with skeletal dysplasia, polycystic kidneys and multiple malformations was first described by Gillessen-Kaesbach et al and subsequently by Nishimura et al. The skeletal features uniformly comprise a round pelvis, mesomelic shortening of the upper limbs and defective ossification of the cervical spine. We studied two unrelated families including three affected fetuses with Gillessen-Kaesbach–Nishimura syndrome using whole-exome and Sanger sequencing, comparative genome hybridization and homozygosity mapping. All affected patients were shown to have a novel homozygous splice variant NM_024740.2: c.1173+2T>A in the ALG9 gene, encoding alpha-1,2-mannosyltransferase, involved in the formation of the lipid-linked oligosaccharide precursor of N-glycosylation. RNA analysis demonstrated skipping of exon 10, leading to shorter RNA. Mass spectrometric analysis showed an increase in monoglycosylated transferrin as compared with control tissues, confirming that this is a congenital disorder of glycosylation (CDG). Only three liveborn children with ALG9-CDG have been previously reported, all with missense variants. All three suffered from intellectual disability, muscular hypotonia, microcephaly and renal cysts, but none had skeletal dysplasia. Our study shows that some pathogenic variants in ALG9 can present as a lethal skeletal dysplasia with visceral malformations as the most severe phenotype. The skeletal features overlap with that previously reported for ALG3- and ALG12-CDG, suggesting that this subset of glycosylation disorders constitutes a new diagnostic group of skeletal dysplasias.