Persistent infection of the cervix with high risk (HR) types of Human Papilloma Virus (HPV) (HR-HPV) can result in precancerous lesions and cancers. However, most HPV infections can be cleared naturally by the immune response without causing disease. Although genetic variations have long been considered as the main explanation for individual heterogeneity in cancer susceptibility, the underlying mechanisms remain unclear. In this project, a panel of routinely taken clinical samples was assessed for 32 rationally selected SNPs with allele frequency related to disease outcome using the Taqman® OpenArray® system. The panel incorporated 475 HR-HPV negative, cytologically-normal cervical samples, 413 HR-HPV positive cervical high grade squamous intraepithelial lesion (HSIL) cases and 62 HR-HPV positive cervical cancers. Two SNPs, rs2234671 and rs2623047, were found with significant differences between HR-HPV negative, cytologically-normal samples and HR-HPV positive cervical HSIL cases. In the validation step, these two SNPs were further genotyped in the same set of samples using TaqMan® SNP genotyping assay and/or LightSNiP assay and in additional samples including 83 HR-HPV positive, cytologically-normal cervical samples, 21 HR-HPV positive cervical cancer cases, 129 HR-HPV positive vulval intraepithelial neoplasia cases and 23 HR-HPV positive vulval cancer cases. Statistical analysis was then performed based on pooled and re-grouped genotyping data of the above-mentioned samples under different genetic models so as to evaluate the associations with different stages in the disease process. After validation, SULF1 rs2623047 revealed a strong significant association with the susceptibility to HR-HPV infection but not with the development of high-grade squamous intraepithelial lesion and the progression to cervical cancer. CXCR1 rs2234671, by contrast, was associated with the progression of HR-HPV-related cancers and the minor allele CXCR1 827C was significantly enriched in HPV16 positive cancers. CXCR1 is a receptor for the chemokine CXCL8/IL-8 and CXCR1 rs2234671 leads to a serine to threonine change in an extracellular loop of the receptor. Functionally, the CXCR1 827C allele was shown to enhance cell motility in response to IL-8 stimulation in a chemotaxis assay with transiently transfected fibroblasts (HEK293 cells) and also in a wound healing assay with stably transduced cervical cancer (CaSki) cells. In addition, significantly increased cell proliferation upon IL-8 treatment was observed in two cervical cancer derived cell lines, CaSki and SiHa, transduced with CXCR1-827C allele, but not in their CXCR1 827G transduced counterparts. These findings suggest that SULF1 rs2623047 and CXCR1 rs2234671 may be genetic risk factors for HR-HPV-related cervical disease and CXCR1 rs2234671 might affect HR-HPV-related cancer susceptibility by functionally altering IL-8-CXCR1 signalling. This information has potential for use in the risk stratification of HR-HPV infected women and may also suggest new therapeutic targets to be exploited for treatment of cervical cancer patients.

Mastitis, or mammary gland inflammation, causes the greatest loss in profit for dairy producers. Mastitis susceptibility differs among cows due to environmental, physiological, and genetic factors. Prior research identified a genetic marker in a chemokine receptor, CXCR1, associated with mastitis susceptibility and decreased neutrophil migration. Current research seeks to identify reasons behind mastitis susceptibility by validating this model through in vivo challenge with Streptococcus uberis and studying specific mechanisms causing impaired neutrophil migration. Holstein cows with GG (n=19), GC (n=28), and CC (n=20) genotypes at CXCR1+777 were challenged intramammarily with S. uberis strain UT888. After challenge 68% of quarters from GG genotype, 74% from CC genotype and only 47% from GC genotype cows had ≥10 colony forming units/ml S. uberis for at least two sampling time points (P<0.05). However, among infected cows, number of S. uberis, somatic cell count, rectal temperature, milk scores and mammary scores were comparable among genotypes throughout infection. These findings suggest that cows with GC genotypes may be more resistant to S. uberis mastitis, but have similar responses if infected. To better understand the mechanisms associated with disease resistance, migration patterns in neutrophils from cows with different CXCR1+777 genotypes were evaluated. Neutrophils from cows with GG (n=11) and CC (n=11) genotypes were isolated and stimulated with zymosan activated sera (ZAS). Cells were fixed and stained for F-actin and evaluated for F-actin content, distribution, and cell morphology. Neutrophils from CC cows had significantly lower average F-actin polymerization than GG cows v (P=0.05). Directed migration of neutrophils from GG (n=10) and CC (n=10) genotypes was imaged and tracking data was analyzed for individual cells. Cells from GG genotype traveled further on an X axis and had higher X/Y movement towards IL8 compared to CC genotype, meaning they moved more directly towards IL8. Our findings suggest lower F-actin polymerization in combination with lower ability to directly move towards IL8 could impair neutrophil response to infection in cows with a CC genotype and may contribute to increased mastitis susceptibility. Finding what makes certain cows more susceptible to mastitis could lead to strategies aimed at improved prevention and treatment of mastitis.

No description available.