Apolipoprotein E Allelic Frequency Altered in Women with Early-onset Breast Cancer

Among women, the most prevalent type of cancer is breast cancer, affecting 1 out of every 8 women in the United States; in Puerto Rico, 70 out of every 100,000 will develop some type of breast cancer. Therefore, a better understand of the potential risk factors for breast cancer could lead to the development of early detection tools. A gene that has been proposed as a risk factor in several populations around the world is Apolipoprotein E (apoE). ApoE functions as a mechanism of transport for lipoproteins and cholesterol throughout the body, with 3 main isoforms present in humans (apoE2, apoE3, and apoE4). Whether or not apoE4 is a risk factor for breast cancer remains controversial. Previous studies have either included test subjects of all ages (20–80) or have focused on late-onset (after age 50) breast cancer; none has concentrated specifically on early-onset (aged 50 and younger) breast cancer. The objectives of this study was to examine (in a Puerto Rican population) the differences in the relative frequency of occurrence of apoE4 in non-breast cancer versus breast cancer patients and to examine, as well, the potential differences of same in early- versus late-onset patients. We found an increased frequency of apoE4 (odds ratio 2.15) only in early-onset breast cancer survivors, which is similar to the findings of those studies that combined or adjusted for age as well as for an association between apoE4 and decreased tumor size. ApoE is also a potential risk factor for long-term cognitive effects after chemotherapy and affects response to hormone replacement. Our data supports the theory that knowing the apoE genotype of women who are at risk of developing breast cancer may be beneficial, as such knowledge would aid in the prediction of tumor size and the development of treatment regimens

Design and Optimization of a Mycoplasma Detection Assay

Mycoplasma are among the smallest free living microorganisms. These bacteria grow slowly, lack a rigid cell wall and are not eliminated by filter sterilization methods used in tissue culture. Mycoplasma infection affects biochemical and genetic aspects of cultured cells, resulting in experimental inconsistency. Therefore, it is necessary to establish routine testing for mycoplasma contamination in tissue culture laboratories. Our goal is to develop a reliable and cost-effective test for mycoplasma in cell culture based on established methods found in literature. We first cloned and sequenced a PCR product from a commercial mycoplasma detection kit. Sequencing revealed the 16s rRNA as the target for mycoplasma detection; we confirmed this target by conducting a literature search. PCR primers were designed using 16s rRNA gene as a target. We set-up reactions and optimized conditions for the real-time PCR assay to detect the target and confirmed amplicon size with agarose gel electrophoresis. We identified that 56oC was the best temperature for the PCR and found that agarose gel electrophoresis was a better detection method because it identified the size to confirm the proper product. The primers we ordered to develop this assay produce the proper band; however, results of several assays have been inconsistent as sometimes a known positive sample fails to amplify. As well, in several PCR reactions the negative showed a signal. The overall reaction needs improvements to have greater reliability and to eliminate all sources of contamination. Research is continuing results are not final

Characterization of HIV-1 RNA forms in the plasma of patients undergoing successful HAART

An assay to characterize plasma human immunodeficiency virus 1 (HIV-1) sequences for patients with low viral loads was developed by combining the selective binding of anti-CD44 MicroBeads with a nested RT-PCR targeting the env C2V4 region. Sequences were obtained from 10 of 20 HIV+ patients who had viral loads below 48 copies/ml. Sequences derived from plasma were compared to those from CD14+ CD16 +monocytes and CD4+ T cells. The plasma sequences were most closely related to those amplified from monocytes, suggesting that during successful antiretroviral therapy, the predominant plasma virus originates from myeloid cells. By characterizing HIV-1 RNA sequences from 8 ml of plasma while avoiding multiple steps, which can lead to contamination and deterioration, this method can help elucidate the viral forms in patients with therapeutically suppressed HIV-1. Understanding the source of residual viremia is crucial in developing approaches for viral eradication