Timo Väisänen

Expression of toll‐like receptor‐9 is increased in poorly differentiated prostate tumors

Toll‐like receptor‐9 (TLR9) is a cellular receptor for bacterial and vertebrate DNA. In addition to cells of the immune system, it is also expressed in various human cancer cell lines, including prostate cancer. We demonstrated previously that synthetic TLR9 ligands induce matrix metalloproteinase‐13‐mediated invasion in TLR9‐expressing prostate cancer cells in vitro. Other studies have suggested possible sex steroid regulation of the function of the various TLRs. The role of TLR9 in the pathophysiology of prostate or any cancer is, however, unknown.

Altered expression of type XIII collagen in keratoconus and scarred human cornea: Increased expression in scarred cornea is associated with myofibroblast transformation.

Purpose: Type XIII collagen (ColXIII) is a transmembrane protein thought to be involved in cell-cell and cell-matrix interactions. We report here on its presence in the normal human cornea and compare the results for keratoconus and scarred corneas. Methods: Immunohistochemistry and in situ hybridization were applied to human corneal samples obtained by penetrating keratoplasty. Results: In the normal human cornea, ColXIII was immunolocalized to the corneal epithelial cells, and to a lesser degree to the stromal keratocytes. The keratoconus cases showed otherwise similar results, but in areas containing Bowman membrane disruptions showed thinned epithelial cells reduced immunostaining for ColXIII, whereas occasionally pronounced immunoreactivity was seen in the stromal keratocytes. The corneal scar samples contained highly increased ColXIII immunostaining by stromal cells in the fibrotic foci, whereas the peripheral areas showed less intense immunostaining. In situ hybridization confirmed that the corneal epithelium and keratocytes actively synthesize the transcript. Immunostaining with &agr;SMA revealed that a substantial proportion of the ColXIII mRNA-expressing cells in the stromal scar tissues was myofibroblasts and that these areas lack CD34 immunoreactivity. Conclusions: The results indicate that ColXIII, which is predominantly confined to the basal corneal cells in the normal cornea, may have a role in the adhesion of corneal epithelial cells to each other and to the underlying basement membrane. Additionally, highly increased expression in scarred corneas suggests that it participates in the corneal wound healing process.

Activated Platelets Provide a Functional Microenvironment for the Antiangiogenic Fragment of Histidine-Rich Glycoprotein

The angiogenesis inhibitor histidine-rich glycoprotein (HRG) constitutes one of several examples of molecules regulating both angiogenesis and hemostasis. The antiangiogenic properties of HRG are mediated via its proteolytically released histidine- and proline-rich (His/Pro-rich) domain. Using a combination of immunohistochemistry and mass spectrometry, we here provide biochemical evidence for the presence of a proteolytic peptide, corresponding to the antiangiogenic domain of HRG, in vivo in human tissue. This finding supports a role for HRG as an endogenous regulator of angiogenesis. Interestingly, the His/Pro-rich peptide bound to the vessel wall in tissue from cancer patients but not to the vasculature in tissue from healthy persons. Moreover, the His/Pro-rich peptide was found in close association with platelets. Relesate from in vitro–activated platelets promoted binding of the His/Pro-rich domain of HRG to endothelial cells, an effect mediated by Zn2+. Previous studies have shown that zinc-dependent binding of the His/Pro-rich domain of HRG to heparan sulfate on endothelial cells is required for inhibition of angiogenesis. We describe a novel mechanism to increase the local concentration and activity of an angiogenesis inhibitor, which may reflect a host response to counteract angiogenesis during pathologic conditions. Our finding that tumor angiogenesis is elevated in HRG-deficient mice supports this conclusion. (Mol Cancer Res 2009;7(11):1792–802)

Modulation of the Cellular Cholesterol Level Affects Shedding of the Type XIII Collagen Ectodomain

Type XIII collagen is a transmembrane protein that also exists as a soluble extracellular variant because of ectodomain shedding by proprotein convertases. Because ectodomain shedding in a growing number of transmembrane proteins has recently been shown to be dependent on their localization in cholesterol-enriched detergent-resistant membrane microdomains, this work aimed at analyzing this aspect of type XIII collagen ectodomain processing. In HT-1080 cells type XIII collagen and its cleaving proprotein convertase furin localized partially in detergent-resistant cholesterol-containing membrane microdomains. Disruption of these domains by lowering either the level or availability of the cellular cholesterol reduced ectodomain shedding, implying that, in such membrane domains correct cholesterol level is important for the regulation of type XIII collagen ectodomain processing. In addition, we show here that ectodomain of type XIII collagen is also shed intracellularly. HT-1080 cells released vesicles from the Golgi apparatus, which contained only the cleaved variant. Intracellular processing and the subsequent entry of the cleaved ectodomain into the vesicles was totally blocked by inhibition of the proprotein convertase function by cell-permeable chloromethylketone, but not with cell-impermeable α1-antitrypsin Portland. This supports the hypothesis of type XIII collagen ectodomain also being cleaved intracellularly in the Golgi and suggests that the intracellular cleavage may act as a gating event in the vesicle-mediated ectodomain secretion.

Collagens XV and XVIII show different expression and localisation in cutaneous squamous cell carcinoma: type XV appears in tumor stroma, while XVIII becomes upregulated in tumor cells and lost from microvessels

As the second most common skin malignancy, cutaneous squamous cell carcinoma (cSCC) is an increasing health concern, while its pathogenesis at molecular level remains largely unknown. We studied the expression and localisation of two homologous basement membrane (BM) collagens, types XV and XVIII, at different stages of cSCC. These collagens are involved in angiogenesis and tumorigenesis, but their role in cancer development is incompletely understood. Quantitative RT‐PCR analysis revealed upregulation of collagen XVIII, but not collagen XV, in primary cSCC cells in comparison with normal human epidermal keratinocytes. In addition, the Ha‐ras‐transformed invasive cell line II‐4 expressed high levels of collagen XVIII mRNA, indicating upregulation in the course of malignant transformation. Immunohistochemical analyses of a large human tissue microarray material showed that collagen XVIII is expressed by tumor cells from grade 1 onwards, while keratinocytes in normal skin and in premalignant lesions showed negative staining for it. Collagen XV appeared instead as deposits in the tumor stroma. Our findings in human cSCCs and in mouse cSCCs from the DMBA‐TPA skin carcinogenesis model showed that collagen XVIII, but not collagen XV or the BM markers collagen IV or laminin, was selectively reduced in the tumor vasculature, and this decrease associated significantly with cancer progression. Our results demonstrate that collagens XV and XVIII are expressed in different sites of cSCC and may contribute in a distinct manner to processes related to cSCC tumorigenesis, identifying these collagens as potential biomarkers in the disease.

Notch Downregulation and Extramedullary Erythrocytosis in Hypoxia-Inducible Factor Prolyl 4-Hydroxylase 2-Deficient Mice

ABSTRACT Erythrocytosis is driven mainly by erythropoietin, which is regulated by hypoxia-inducible factor (HIF). Mutations in HIF prolyl 4-hydroxylase 2 (HIF-P4H-2) (PHD2/EGLN1), the major downregulator of HIFα subunits, are found in familiar erythrocytosis, and large-spectrum conditional inactivation of HIF-P4H-2 in mice leads to severe erythrocytosis. Although bone marrow is the primary site for erythropoiesis, spleen remains capable of extramedullary erythropoiesis. We studied HIF-P4H-2-deficient (Hif-p4h-2gt/gt) mice, which show slightly induced erythropoiesis upon aging despite nonincreased erythropoietin levels, and identified spleen as the site of extramedullary erythropoiesis. Splenic hematopoietic stem cells (HSCs) of these mice exhibited increased erythroid burst-forming unit (BFU-E) growth, and the mice were protected against anemia. HIF-1α and HIF-2α were stabilized in the spleens, while the Notch ligand genes Jag1, Jag2, and Dll1 and target Hes1 became downregulated upon aging HIF-2α dependently. Inhibition of Notch signaling in wild-type spleen HSCs phenocopied the increased BFU-E growth. HIFα stabilization can thus mediate non-erythropoietin-driven splenic erythropoiesis via altered Notch signaling.

Compound traditional serrated adenoma and sessile serrated adenoma

For many years, it was understood that the vast majority of the colorectal carcinomas in the general population evolved from conventional adenomas via the adenoma–carcinoma sequence. In 1990, Longacre and Fenoglio-Preiser first described polyps characterised by serrated architecture and unequivocal dysplasia1; these neoplastic polyps were subsequently called traditional serrated adenomas (TSAs).2 More recently, serrated colorectal polyps—hyperplastic polyps, sessile serrated adenoma/polyps (SSA/P)3 and TSAs3 ,4—have emerged as an alternative pathway of colorectal carcinogenesis. It has been estimated that in the general population about 30% of the CRC progress via the serrated pathway.3 So far, the histogenesis and natural history of TSAs have remained undisclosed. A 73-year-old woman consulted for gastrointestinal bleeding. The patient had alopecia, nail atrophy, iron deficiency and anaemia. Colonoscopic and gastroscopic examinations disclosed multiple colorectal polyps and confluent gastroduodenal polyps, respectively. Histology showed to be cystic hamartomas. MRI showed no jejunal or ileum polyps. She had no chronic diarrhoea, protein-losing enteropathy, family history of polyposis or of colorectal cancer. Molecular genetic testing disclosed no SMAD4 or BMPR1A genes mutations. Because of the dense colorectal polyposis and repeated bleedings, a subtotal colectomy was carried out under a diagnosis of Cronkhite-Canada syndrome (CCS). The …