Cystatin C Deficiency Promotes Epidermal Dysplasia in K14-HPV16 Transgenic Mice

Cysteine protease cathepsins are important in extracellular matrix protein degradation, cell apoptosis, and angiogenesis. Mice lacking cathepsins are protected from tumor progression in several animal models, suggesting that the regulation of cathepsin activities controls the growth of various malignant tumors.We tested the role of cathepsins using a mouse model of multistage epithelial carcinogenesis, in which the human keratin-14 promoter/enhancer drove the expression of human papillomavirus type 16 (HPV16) early region E6/E7 transgenes. During the progression of premalignant dysplasia, we observed increased expression of cysteine protease cathepsin S, but concomitantly reduced expression of cathepsin endogenous inhibitor cystatin C in the skin tissue extract. Absence of cystatin C in these transgenic mice resulted in more progression of dysplasia to carcinoma in situ on the face, ear, chest, and tail. Chest and ear skin extract real time PCR and immunoblot analysis, mouse serum sample ELISA, tissue immunohistological analysis, and tissue extract-mediated in vitro elastinolysis and collagenolysis assays demonstrated that cystatin C deficiency significantly increased cathepsin expression and activity. In skin from both the chest and ear, we found that the absence of cystatin C reduced epithelial cell apoptosis but increased proliferation. From the same tissue preparations, we detected significantly higher levels of pro-angiogenic laminin 5-derived γ2 peptides and concurrently increased neovascularization in cystatin C-deficient mice, compared to those from wild-type control mice.Enhanced cathepsin expression and activity in cystatin C-deficient mice contributed to the progression of dysplasia by altering premalignant tissue epithelial proliferation, apoptosis, and neovascularization

Rh-I-UEA-1 Polymerized Liposomes Target and Image Adenomatous Polyps in the APC

Mutated adenomatous polyposis coli (APC) genes predispose transformations to neoplasia progressing to colorectal carcinoma (CRC). Early detection facilitates clinical management and therapy. Novel lectin-mediated polymerized targeted liposomes (Rh-I-UEA-1), with polyp specificity and incorporated imaging agents, were fabricated to locate and image adenomatous polyps in APC(Min/+) mice. The biomarker α-L-fucose covalently joins liposomal conjugated lectin ulex euroapeus agglutinin (UEA-1), via glycosidic linkage to the polyp mucin layer. Multispectral optical imaging (MSI) corroborated a global perspective of specific binding (Rhodamine B 532 nm emission, 590–620 nm excitation) of targeted Rh-I-UEA-1 polymerized liposomes to polyps with 1.4× fold labeling efficiency. High-resolution co-registered optical coherence tomography (OCT) and fluorescence molecular imaging (FMI) reveal spatial correlation of contrast distribution and tissue morphology. Freshly excised APC(Min) bowels were incubated with targeted liposomes (UEA-1 lectin), control liposomes (no lectin), or Omnipaque and imaged by the three techniques. CT quantitative analyses did not confirm targeted liposomes more strongly bound polyps than nontargeted liposomes or Omnipaque alone. OCT, with anatomical depth capabilities, along with the co-registered FMI, substantiated Rh-I-UEA-1 liposome binding along the mucinous polyp surface. UEA-1 lectin denotes α-L-fucose biomarker carbohydrate expression at the mucin glycoprotein layer; Rh-I-UEA-1 polymerized liposomes target and image adenomatous polyps in APC(Min) mice

A homozygous founder missense variant in arylsulfatase G abolishes its enzymatic activity causing atypical Usher syndrome in humans.

We aimed to identify the cause of disease in patients suffering from a distinctive, atypical form of Usher syndrome. Whole-exome and genome sequencing were performed in five patients from three families of Yemenite Jewish origin, suffering from distinctive retinal degeneration phenotype and sensorineural hearing loss. Functional analysis of the wild-type and mutant proteins was performed in human fibrosarcoma cells. We identified a homozygous founder missense variant, c.133G>T (p.D45Y) in arylsulfatase G (ARSG). All patients shared a distinctive retinal phenotype with ring-shaped atrophy along the arcades engirdling the fovea, resulting in ring scotoma. In addition, patients developed moderate to severe sensorineural hearing loss. Both vision and hearing loss appeared around the age of 40 years. The identified variant affected a fully conserved amino acid that is part of the catalytic site of the enzyme. Functional analysis of the wild-type and mutant proteins showed no basal activity of p.D45Y. Homozygosity for ARSG-p.D45Y in humans leads to protein dysfunction, causing an atypical combination of late-onset Usher syndrome. Although there is no evidence for generalized clinical manifestations of lysosomal storage diseases in this set of patients, we cannot rule out the possibility that mild and late-onset symptoms may appear