A pilot study to evaluate the changes in venous blood gas parameters and hypoxia biomarkers in health care workers using different kinds of masks.

Background: The study is aimed to investigate the metabolic alterations and changes in biochemical parameters associated with extended mask. Methods: It was a prospective comparative study conducted on 129 participants comprised of 37 healthy controls and 92 health care workers using different kind of masks like, cloth mask, surgical masks and N95-FFR/PPE. Two samples on day-1 and day-10 were collected for analysis of blood gas parameters, serum hypoxia-inducible factor-α (HIF-α), and erythropoietin (EPO). Results: Oxygen saturation percentage (sO2) of 72.68 (P = 0.033) was significantly low, whereas, Na+ (P = 0.05) and Ca2+ (P < 0.001) were raised in exposed individuals than the healthy controls. The serum HIF-α level of 3.26 ng/mL, was considerable higher in the exposed individuals than controls (P = 0.001). pO2 and sO2 were the lowest and HIF-α and EPO were raised in N95-FFR/PPE of all mask users (P < 0.01). A significant difference was evidenced for pCO2, pH, Na+, Ca2+, and EPO in the exposed group. A positive correlation between the duration of mask use (in hours) with HIF-α (r = 0.247, P = 0.005) and Ca2+ (r = 0.306, P < 0.001) was observed. The major complaints in N95-FFR/PPE users were headache (15.2%) and polydipsia (33.3%). Conclusion: The study findings depicted a significant metabolic alterations in PPE/N95 users which could be due to chronic hypoxic exposure of the tissues.

Alcohol metabolism in human cells causes DNA damage and activates the Fanconi anemia-breast cancer susceptibility (FA-BRCA) DNA damage response network.

BACKGROUND: We recently reported that exposure of human cells in vitro to acetaldehyde resulted in the activation of the Fanconi anemia-breast cancer susceptibility (FA-BRCA) DNA damage response network. METHODS: To determine whether intracellular generation of acetaldehyde from ethanol metabolism can cause DNA damage and activate the FA-BRCA network, we engineered HeLa cells to metabolize alcohol by expression of human alcohol dehydrogenase (ADH) 1B. RESULTS: Incubation of HeLa-ADH1B cells with ethanol (20 mM) resulted in acetaldehyde accumulation in the media, which was prevented by co-incubation with 4-methyl pyrazole (4-MP), a specific inhibitor of ADH. Ethanol treatment of HeLa-ADH1B cells produced a 4-fold increase in the acetaldehyde-DNA adduct and N(2)-ethylidene-dGuo and also resulted in the activation of the FA-BRCA DNA damage response network, as indicated by a monoubiquitination of FANCD2 and phosphorylation of BRCA1. Ser 1524 was identified as 1 site of BRCA1 phosphorylation. The increased levels of DNA adducts, FANCD2 monoubiquitination, and BRCA1 phosphorylation were all blocked by 4-MP, indicating that acetaldehyde, rather than ethanol itself, was responsible for all 3 responses. Importantly, the ethanol concentration we used is within the range that can be attained in the human body during social drinking. CONCLUSIONS: Our results indicate that intracellular metabolism of ethanol to acetaldehyde results in DNA damage, which activates the FA-BRCA DNA damage response network.

Molecular constitution of breast but not other reproductive tissues is rich in growth promoting molecules: a possible link to highest incidence of tumor growths.

In the current study we tested if highest incidence of benign as well as cancer growths in breast tissue is due to constitutive molecular composition of this tissue. To delineate the molecular basis, we compared the expression of nine functional gene modules (total 578 genes) that regulate major positive growth and negative inhibitory signals in normal breast with two other reproductive tissues, ovary and uterus. We present data to demonstrate that breast tissues constitutively have very highly elevated levels of several growth promoting molecules and diminished levels of inhibitory molecules which may, in part, contribute for highest incidence of tumor growths in this tissue.

Molecular risk assessment for breast cancer development in patients with ductal hyperplasias.

PURPOSE: It has been reported that approximately a million women are diagnosed with benign breast lesions that include ductal hyperplasias per year in the United States. Recent studies that followed women with benign lesions have established that about 8% to 9% of them will subsequently develop invasive breast cancer (IBC). However, currently, there are no means of identifying a subclass of "true precancerous tissues" in women with ductal hyperplasias who will subsequently develop cancer. The purpose of this study is to investigate whether expression of hyaluronoglucosaminidase 1 (HYAL1), a known tumor promoter, in hyperplastic tissues identifies a "true precancerous stage" and predicts subsequent IBC development. EXPERIMENTAL DESIGN: A retrospective study was conducted with archival benign tissues of various histologic types and clinical information on development/nondevelopment of IBC. The control group was hyperplastic tissues from women who had no prior history of IBC and did not develop cancer in 5 to 7 years after diagnosis (n = 81). The test group was hyperplastic tissues from patients who developed cancer (n = 82). HYAL1 expression was studied by immunohistochemistry, and the results were statistically analyzed for significant association to develop cancer (P value), specificity, sensitivity, positive predictive value, and negative predictive value. RESULTS: Statistical analysis of HYAL1 expression data showed very highly significant association between its expression and subsequent cancer development (P = 0) and very high sensitivity (0.83), specificity (0.84), positive predictive value (0.84), and negative predictive value (0.83). CONCLUSIONS: The expression of HYAL1 in ductal hyperplastic tissues is a strong predictor of subsequent development of IBC; therefore, it can be applied as a diagnostic marker either singly or in combination with other marker(s) to screen benign tissues to predict subsequent development of IBC. Detection at the precancerous stage and treatment could drastically cut down breast cancer incidence and deaths from it.

The Cell Surface Estrogen Receptor, G Protein- Coupled Receptor 30 (GPR30), is Markedly Down Regulated During Breast Tumorigenesis.

BACKGROUND: GPR30 is a cell surface estrogen receptor that has been shown to mediate a number of non-genomic rapid effects of estrogen and appear to balance the signaling of estrogen and growth factors. In addition, progestins appear to use GPR30 for their actions. Therefore, GPR30 could play a critical role in hormonal regulation of breast epithelial cell integrity. Deregulation of the events mediated by GPR30 could contribute to tumorigenesis. METHODS: To understand the role of GPR30 in the deregulation of estrogen signaling processes during breast carcinogenesis, we have undertaken this study to investigate its expression at mRNA levels in tumor tissues and their matched normal tissues. We compared its expression at mRNA levels by RT quantitative real-time PCR relative to GAPDH in ERα"-positive (n = 54) and ERα"-negative (n = 45) breast cancer tissues to their matched normal tissues. RESULTS: We report here, for the first time, that GPR30 mRNA levels were significantly down-regulated in cancer tissues in comparison with their matched normal tissues (p < 0.0001 by two sided paired t-test). The GPR30 expression levels were significantly lower in tumor tissues from patients (n = 29) who had lymph node metastasis in comparison with tumors from patients (n = 53) who were negative for lymph node metastasis (two sample t-test, p < 0.02), but no association was found with ERα, PR and other tumor characteristics. CONCLUSIONS: Down-regulation of GPR30 could contribute to breast tumorigenesis and lymph node metastasis.

Estrogen receptors beta4 and beta5 are full length functionally distinct ERbeta isoforms: cloning from human ovary and functional characterization.

We describe here the cloning and functional characterization of two unique ER isoforms, ERbeta4 and ERbeta5. The full length ERbeta4 and ERbeta5 were identified by asymmetric PCR using human ovary cDNA, cloning, and sequence analyses. Both receptors share identical sequences with ERbeta1 from exon 1 to exon 7. In the place of exon 8, ERbeta4 has unique sequences arising from a region downstream of the ERbeta gene and upstream of the SYNE2 gene. ERbeta5 has sequences arising from retention of the 5' end of the intron between exon 7 and 8. Both receptors bind promoter sequences on DNA but do not bind estrogen. They translocate to the nucleus and exhibit three to four times higher estrogen-independent transcriptional activity than ERbeta1. When co-transfected with ERalpha, they predominantly form heterodimers and negatively regulate its transcriptional activity. Estrogen-independent transcriptional activity of ERbeta5, but not ERbeta4, was inhibited by ERalpha, demonstrating for the first time that ERalpha regulates ERbeta. Tissue-specific expression of ERbeta4 and ERbeta5, together with their ligand-independent transcriptional properties and ERalpha modulating activities, could have a number of implications in seemingly unlinked biological processes regulated by estrogen.

Estrogen receptor alpha-negative breast cancer tissues express significant levels of estrogen-independent transcription factors, ERbeta1 and ERbeta5: potential molecular targets for chemoprevention.

We have investigated the expression of two estrogen receptor beta (ERbeta) isoforms, ERbeta1 and ERbeta5, which activate gene transcription independent of estrogen or growth factors, in ERalpha-negative breast cancer tissues. We report here, for the first time, that ERalpha-negative tissues express significant levels of ERbeta1 and ERbeta5, and their expression levels are not different from levels in ERalpha positive tumors. However, significant differences exist between the two racial groups, African American and Caucasian, in that the patients from the former group express higher levels of ERbeta1 and ERbeta5 but not ERalpha. These two transcription factors could be potential molecular targets for designing chemopreventive drugs to treat ERalpha-negative breast cancers.

Identification of MMP-1 as a putative breast cancer predictive marker by global gene expression analysis.

Breast cancer is the second leading cause of cancer death for women in the United States. In 2005, about 215,000 cases of invasive breast cancer (IBC) and 50,000 cases of ductal carcinoma in situ will be diagnosed and 40,000 women will die of IBC in the US. Yet there is presently no molecular marker that can be used to detect a precancerous state or identify which premalignant lesions will develop into invasive breast cancer. Here we report the gene expression analysis of atypical ductal hyperplastic tissues from patients with and without a history of breast cancer. We identify MMP-1 as a candidate marker that may be useful for identification of breast lesions that can develop into cancer.

Correlation of expression of BP1, a homeobox gene, with estrogen receptor status in breast cancer.

BACKGROUND: BP1 is a novel homeobox gene cloned in our laboratory. Our previous studies in leukemia demonstrated that BP1 has oncogenic properties, including as a modulator of cell survival. Here BP1 expression was examined in breast cancer, and the relationship between BP1 expression and clinicopathological data was determined. METHODS: Total RNA was isolated from cell lines, tumors, and matched normal adjacent tissue or tissue from autopsy. Reverse transcription polymerase chain reaction was performed to evaluate BP1 expression. Statistical analysis was accomplished with SAS. RESULTS: Analysis of 46 invasive ductal breast tumors demonstrated BP1 expression in 80% of them, compared with a lack of expression in six normal breast tissues and low-level expression in one normal breast tissue. Remarkably, 100% of tumors that were negative for the estrogen receptor (ER) were BP1-positive, whereas 73% of ER-positive tumors expressed BP1 (P = 0.03). BP1 expression was also associated with race: 89% of the tumors of African American women were BP1-positive, whereas 57% of those from Caucasian women expressed BP1 (P = 0.04). However, there was no significant difference in BP1 expression between grades I, II, and III tumors. Interestingly, BP1 mRNA expression was correlated with the ability of malignant cell lines to cause breast cancer in mice. CONCLUSION: Because BP1 is expressed abnormally in breast tumors, it could provide a useful target for therapy, particularly in patients with ER-negative tumors. The frequent expression of BP1 in all tumor grades suggests that activation of BP1 is an early event.

Estrogen receptor beta splice variant mRNAs are differentially altered during breast carcinogenesis.

We previously identified 10 exon deletion ERbeta variant mRNAs in various human tissues [FEBS Lett. 516 (2002) 133]. In the current study, we have investigated the expression of these variant mRNAs in normal breast tissues and their alterations in cancer tissues. A total of 43 cancer tissues in comparison with their matched normal tissues were analyzed by RT PCR using the newly developed 'Splice Targeted Primer Approach'. The data presented here show that normal breast tissues express 9 of the 10 identified variant mRNAs. Of the nine variants, the mRNAs with exons 5-6 deletions were significantly decreased ( approximately 80%) in a large majority of cancer tissues (two-sided paired t-test, n=43 patients, P<0.00001). The expression of ERbeta exon 5Delta, that could potentially have transactivating property in the absence of hormone, was changed differently among different grade tumors (analysis of variance F-test, n=43 patients, P=0.0452; Kruskal-Wallis test, n=43 patients, P=0.0356). When change in expression of ERbeta exon 5Delta mRNA levels was used as a categorical variable, a significant association was found between the change status (increase, no change, decrease) of this variant and grade of the tumor (Fischer's exact test, n=43 patients, P=0.0129). In particular, it was significantly increased in grade III tumors and decreased in grade II tumors. This variant was also changed differently in pre- and post-menopausal women. Its expression levels were increased in the tumors of post-menopausal women (mean change=3.6685), while they were decreased in pre-menopausal women (mean change=-24.3662). Thus a significant association was observed between the expression of this variant and menopausal status (a two-sided paired t-test, n=43 patients, P=0.03). Other variants were either expressed at very low frequency or not significantly altered.

Identification of ten exon deleted ERbeta mRNAs in human ovary, breast, uterus and bone tissues: alternate splicing pattern of estrogen receptor beta mRNA is distinct from that of estrogen receptor alpha.

Four different human tissues and breast cancer cell lines were screened to identify exon deletion variant transcripts of estrogen receptor beta (ERbeta) by reverse transcription-polymerase chain reaction using the 'splice targeted primer approach' that amplifies each category of exon deleted variants as a separate gene population. A total of 10 different variant mRNAs that have deletions in various combination of exons were identified by sequence analysis. They were exon 2Delta; exons 2 and 5-6Delta; exon 3Delta; exon 4Delta; exon 5Delta; exons 5 and 2Delta; exon 6Delta; exons 6 and 2Delta; exons 6, 2-3Delta; and exons 5-6Delta. In some cases, deletion of an exon appears to be associated with a mutation of a specific base. Although ERalpha and ERbeta are highly homologous, have identical exon and functional domain organization, exhibit similar ligand-binding profiles and interact with identical DNA response elements, the sequence of exon skipping in ERbeta pre-mRNA appears to be distinct from that of ERalpha mRNA. Furthermore, results described here also suggest that alternate splicing of ERbeta mRNA is tissue specific. The presence of a ERbeta variant profile together with other ER isoforms in a tissue may have functional implications in binding and response to a particular ligand.