Time-Restricted Feeding Shifts the Skin Circadian Clock and Alters UVB-Induced DNA Damage.

The epidermis is a highly regenerative barrier protecting organisms from environmental insults, including UV radiation, the main cause of skin cancer and skin aging. Here, we show that time-restricted feeding (RF) shifts the phase and alters the amplitude of the skin circadian clock and affects the expression of approximately 10% of the skin transcriptome. Furthermore, a large number of skin-expressed genes are acutely regulated by food intake. Although the circadian clock is required for daily rhythms in DNA synthesis in epidermal progenitor cells, RF-induced shifts in clock phase do not alter the phase of DNA synthesis. However, RF alters both diurnal sensitivity to UVB-induced DNA damage and expression of the key DNA repair gene, Xpa. Together, our findings indicate regulation of skin function by time of feeding and emphasize a link between circadian rhythm, food intake, and skin health.

The circadian clock in skin: implications for adult stem cells, tissue regeneration, cancer, aging, and immunity.

Historically, work on peripheral circadian clocks has been focused on organs and tissues that have prominent metabolic functions, such as the liver, fat, and muscle. In recent years, skin has emerged as a model for studying circadian clock regulation of cell proliferation, stem cell functions, tissue regeneration, aging, and carcinogenesis. Morphologically, skin is complex, containing multiple cell types and structures, and there is evidence for a functional circadian clock in most, if not all, of its cell types. Despite the complexity, skin stem cell populations are well defined, experimentally tractable, and exhibit prominent daily cell proliferation cycles. Hair follicle stem cells also participate in recurrent, long-lasting cycles of regeneration: the hair growth cycles. Among other advantages of skin is a broad repertoire of available genetic tools enabling the creation of cell type-specific circadian mutants. Also, due to the accessibility of skin, in vivo imaging techniques can be readily applied to study the circadian clock and its outputs in real time, even at the single-cell level. Skin provides the first line of defense against many environmental and stress factors that exhibit dramatic diurnal variations such as solar ultraviolet (UV) radiation and temperature. Studies have already linked the circadian clock to the control of UVB-induced DNA damage and skin cancers. Due to the important role that skin plays in the defense against microorganisms, it also represents a promising model system to further explore the role of the clock in the regulation of the body's immune functions. To that end, recent studies have already linked the circadian clock to psoriasis, one of the most common immune-mediated skin disorders. Skin also provides opportunities to interrogate the clock regulation of tissue metabolism in the context of stem cells and regeneration. Furthermore, many animal species feature prominent seasonal hair molt cycles, offering an attractive model for investigating the role of the clock in seasonal organismal behaviors.

Risk factors associated with the diagnosis of abdominal aortic aneurysm in patients screened at a regional Veterans Affairs health care system.

BACKGROUND: An active abdominal aortic aneurysm (AAA) screening program at a regional Veterans Affairs (VA) health system identifies patients at risk for AAA. The purpose of this study is to evaluate unique risk factors associated with the AAA diagnosis upon AAA screening examination to identify the most at risk patients for AAA. METHODS: Data were extracted from a regional VA health care system to identify patients who underwent AAA screening within a 3-year period. An aortic diameter ≥3.0 cm was defined as an AAA. Patient risk factors included age, body mass index, total cholesterol, estimated glomerular filtration rate (eGFR), statin use, and active smoking status; the presence of hypertension, diabetes, coronary artery disease (CAD), chronic obstructive pulmonary disease (COPD), or peripheral vascular disease (PVD) was also evaluated. Risk factors were compared in a multivariate analysis between patients with AAA and patients with a normal aorta. RESULTS: A total of 6,142 patients (mean ± SD age: 72.7 ± 5.3 years) were screened for AAA between January 2007 and December 2009. A total of 469 patients (7.6%) with AAA were identified. The following risk factors were significantly associated with a diagnosis of AAA: age >75 years (39.6% vs. 28.9%; P < 0.001), prevalence of CAD (43.1% vs. 28.5%; P < 0.001), COPD (26% vs. 11.4%; P < 0.001), PVD (37.3% vs. 7.7%; P < 0.001), eGFR <60 mL/min (36.7% vs. 24.3%; P < 0.001), and current smoking (23.2% vs. 15.3%; P < 0.001). The risk factors significantly associated with normal aortic size were the presence of diabetes (18.6% vs. 27.4%; P < 0.001) and total cholesterol ≥200 mg/dL (10.4% vs. 15%; P = 0.04). CONCLUSIONS: The diagnosis of AAA in a large screening study is typically identified in patients who are at high risk for cardiovascular disease. The presence of diabetes is a major cardiovascular risk factor that is more associated with normal aorta when compared to patients with the AAA diagnosis. Total cholesterol ≥200 mg/dL was associated with decreased AAA risk, and renal insufficiency was associated with increased AAA risk.

Increased levels of CD34+ cells are associated in patients with abdominal aortic aneurysms compared with patients with peripheral vascular disease.

BACKGROUND: Circulating progenitor cells are integral to vascular health and effectively predict vascular reactivity. CD34 is a known marker of circulating progenitor cells. Few studies have examined the role of CD34+ cells in abdominal aortic aneurysm (AAA) disease and peripheral vascular disease (PVD). The aim of this study was to compare the percentage of CD34+ cells between patients with AAA versus PVD. MATERIALS AND METHODS: We collected peripheral whole blood from AAA or PVD patients. The blood was stained with fluorescently labeled antibodies against CD34 or isotype controls. We collected data using a flow cytometer and analyzed them. We also recorded risk factors such as hypertension, diabetes, total cholesterol, serum white blood cells, serum creatinine, body mass index, blood pressure, statin use, current smoking status, coronary artery disease, cerebral vascular accident, and chronic obstructive pulmonary disease. RESULTS: We enrolled 24 patients in this study (AAA, n = 12; PVD, n = 12). The AAA patients had a greater percentage of CD34+ cells compared with PVD patients. (r = 0.84; P = 0.016). There were no significant risk factors differences between AAA and PVD patients. CONCLUSIONS: Based on CD34+ cell counts, AAA is a less severe vascular disease than PVD. Whether CD34+ cells can serve as a biomarker for risk stratification or a potential therapy warrants further study.

Outcomes of an abdominal aortic aneurysm screening program.

OBJECTIVE: In 2007, Medicare guidelines were established to identify persons at risk for the presence of an abdominal aortic aneurysm (AAA). The purpose of this study is to evaluate the 5-year outcomes of an AAA screening program in a regional Veterans Affairs (VA) health care system. METHODS: Data were extracted from a regional VA health care network identifying all veteran males 65 to 75 years of age who smoked at least 100 cigarettes during their lifetime. In 2007, an AAA screening mandate was implemented allowing patients meeting screening criteria to be evaluated for AAA as part of the patient's health maintenance. AAA is identified as an aortic diameter size of 3.0 cm or greater. Clinician adherence to screening protocols and referral to a vascular surgeon for aneurysms >5.5 cm were also evaluated. RESULTS: A total of 9751 patients (71.5 ± 5.6 standard deviation years of age) were screened for an AAA over a 5-year period from January 1, 2007 to December 31, 2011. A total of 698 aneurysms (7.1%) were found. Referrals to a vascular surgeon were made on 45 patients with aneurysms >5.5 cm. Over a 5-year period, a total of 2754 patients (28.2%) were inappropriately screened: 416 patients were under 65 years old, 2243 patients were over 75 years old, 36 patients were women, and 123 patients without aneurysms had multiple screenings. In 2007, during the first year of implementation, 39.2% of patients were inappropriately screened. Over the next 4 years, inappropriate screenings decreased with 33.7% in 2008, 28.6% in 2009, 17.7% in 2010, and 14.3% in 2011. CONCLUSIONS: A large AAA screening program at the VA detects more aneurysms, but at smaller diameters than that published in clinical trials. Over time, the number of inappropriate AAA screenings has continued to decrease, demonstrating greater awareness and application of the AAA screening guidelines by primary care providers. Developing surveillance guidelines for small and medium aneurysms is a potential area for future research.

Monocytic adhesion molecule expression and monocyte-endothelial cell dysfunction are increased in patients with peripheral vascular disease versus patients with abdominal aortic aneurysms.

BACKGROUND: Statin therapy is used in the medical management of patients with peripheral vascular disease (PVD) and abdominal aortic aneurysm (AAA) for the pleiotropic and anti-inflammatory benefits. We hypothesize that the inflammatory mechanisms of monocyte-endothelial cell interactions in endothelial barrier dysfunction are more significant in patients with PVD compared with those with AAA. The purpose of this study was to assess patient peripheral blood monocyte adhesion molecules by flow cytometry and monocyte-induced endothelial barrier dysfunction by using an in vitro endothelial cell layer and electric cell-substrate impedance sensing (ECIS) system. METHODS: Peripheral blood was collected from patients with either PVD (ankle-brachial index <0.9, toe-arm index <0.8, or required lower extremity vascular intervention) or AAA (aortic diameter >3.0 cm). Monocytes were isolated from fresh whole blood using an accuspin-histopaque technique. The separated monocytes underwent flow cytometry analysis to evaluate the expression levels of the cell membrane adhesion molecules: CD18, CD11a/b/c, and very late antigen-4. Endothelial cell function was assessed by adding monocytes to an endothelial monolayer on ECIS arrays and coculturing overnight. Peak changes in transendothelial electrical resistance were measured and compared between patient groups. RESULTS: Twenty-eight monocyte samples were analyzed for adhesion molecules (PVD, 19 and AAA, 9) via flow cytometry, and 11 patients were evaluated for endothelial dysfunction (PVD, 7 and AAA, 4) via ECIS. There was no significant difference between risk factors among PVD and AAA patients except for age, where AAA patients were significantly older than PVD patients in both flow cytometry and ECIS groups (P=0.02 and 0.01, respectively). There were significantly higher levels of adhesion molecules CD11a, CD18, and CD11c (averaged mean fluorescent intensity P values: 0.047, 0.038, and 0.014, respectively) in PVD patients compared with AAA patients. No significant difference was found for CD11b and very late antigen-4 expression (P=0.21 and 0.15, respectively). There was significantly more monocyte-endothelial cell dysfunction in patients with PVD versus patients with AAA, with a maximal effect seen at 15h after monocyte addition (P=0.032). CONCLUSIONS: Patients with PVD have increased expression levels of certain monocyte adhesion molecules and greater monocyte-induced endothelial layer dysfunction compared with those with AAA. This may lead to other methods of targeted therapy to improve outcomes of these vascular patients.

Dual EGFR/HER2 inhibition sensitizes prostate cancer cells to androgen withdrawal by suppressing ErbB3.

PURPOSE: Patients with recurrent prostate cancer are commonly treated with androgen withdrawal therapy (AWT); however, almost all patients eventually progress to castration resistant prostate cancer (CRPC), indicating failure of AWT to eliminate androgen-sensitive prostate cancer. The overall goal of these studies is to determine whether dual inhibition of the receptor tyrosine kinases epidermal growth factor receptor (EGFR) and HER2 would prolong the effectiveness of this treatment in prostate cancer. EXPERIMENTAL DESIGN: We used androgen-dependent LNCaP cells and its CRPC sublines LNCaP-AI and C4-2. Additional data were collected in pRNS-1-1 cells stably expressing a mutant androgen receptor (AR-T877A), and in nude mice harboring CWR22 tumors. Studies utilized EGFR inhibitors erlotinib and AG1478, and HER2 inhibitors trastuzumab and AG879. RESULTS: Dual EGFR/HER2 inhibition induced apoptosis selectively in androgen-sensitive prostate cancer cells undergoing AWT, but not in the presence of androgens, or in CRPC cells. We show that AWT alone failed to induce significant apoptosis in androgen-dependent cells, due to AWT-induced increase in HER2 and ErbB3, which promoted survival by increasing Akt phosphorylation. AWT-induced ErbB3 stabilized the AR and stimulated PSA, while it was inactivated only by inhibition of both its dimerization partners EGFR and HER2 (prostate cancer cells do not express ErbB4); but not the inhibition of any one receptor alone, explaining the success of dual EGFR/HER2 inhibition in sensitizing androgen-dependent cells to AWT. The effectiveness of the inhibitors in suppressing growth correlated with its ability to prevent Akt phosphorylation. CONCLUSION: These studies indicate that dual EGFR/HER2 inhibition, administered together with AWT, sensitize prostate cancer cells to apoptosis during AWT.