Expression of a cloned denV gene of bacteriophage T4 in Escherichia coli.

A 713-base-pair Hae III fragment from bacteriophage T4 encompassing the denV gene with its preceding promoter has been cloned in a pBR322-derived positive-selection vector and introduced into a variety of DNA repair-deficient uvr and rec and uvr,rec Escherichia coli strains. The denV gene was found to be expressed, probably from its own promoter, causing pyrimidine dimer incision-deficient uvrA, uvrB, uvrC strains to be rescued by the denV gene. A uvrD (DNA helicase II) strain was also complemented, but to a lesser extent. A wild-type strain did not seem to be affected at the UV doses tested. Surprisingly, all recA, recB, and recC strains tested also showed an increased UV resistance, perhaps by reinforcement of the intact uvr system in these strains. Complementation of denV- T4 strains and host-cell reactivation of lambda phage was also observed in denV+ E. coli strains. Equilibrium sedimentation showed that DNA repair synthesis occurred in a UV-irradiated uvrA E. coli strain carrying the cloned denV gene. Southern blotting confirmed our earlier results [Valerie, K., Henderson, E. E. & de Riel, J. K. (1984) Nucleic Acids Res. 12, 8085-8096] that the denV gene is located at 64 kilobases on the T4 map. Phage T2 (denV-) did not hybridize to a denV-specific probe.     

Genetic complementation of UV-induced DNA repair in Chinese hamster ovary cells by the denV gene of phage T4.

The denV gene of phage T4, encoding the pyrimidine dimer-specific DNA repair enzyme endonuclease V, has been introduced by DNA transfection into the UV-sensitive DNA repair-deficient Chinese hamster ovary (CHO) cell line UV5. Transformants were first selected for resistance to the antibiotic G418 conferred by the neo gene from Tn5 carried by the same plasmid. A majority of the isolated G418-resistant UV5 clones also showed an increased resistance to 254-nm UV light. One clone, designated I-A1, was found to have an intermediate level of colony-forming ability after UV irradiation when compared to UV5 and wild-type AA8 cells. A Southern blot showed that I-A1 carries a single integrated intact copy of the denV gene. Alkaline sucrose gradients revealed a dose-dependent appearance of breaks in the DNA of I-A1 cells following UV-irradiation, while unirradiated cells did not exhibit any significant breaks. Analysis of DNA repair by isopycnic sedimentation showed that DNA excision repair by I-A1 was at least equal to the level of repair in AA8 cells. These results show that the prokaryotic denV gene can restore UV repair capabilities in vivo to CHO UV5 cells defective in repair of UV-induced damage.     

Extent of Primary Breast Cancer Surgery: Standards and Individualized Concepts

Surgery is still a main therapeutic option in breast cancer treatment. Nowadays, methods of resection and reconstruction vary according to different tumors and patients. This review presents and discusses standards of care and arising questions on how radical primary breast cancer surgery should be according to different clinical situations. In most early breast cancer patients, breast conservation is the method of choice. The discussion on resection margins is still controversial as different studies show conflicting results. Modified radical mastectomy is the standard in locally advanced breast cancer patients, although there are different promising approaches to spare skin or even the nipple-areola complex. A sentinel node biopsy is the standard of care in clinically node-negative invasive breast cancer patients, whereas the significance of axillary lymphonodectomy seems to be questioned through a number of different findings. Although there are interesting findings to modify surgical approaches in very young or elderly breast cancer patients, it will always be an individualized approach if we do not adhere to current guidelines. Up to date, there are no special surgical procedures in BRCA mutation carriers or patients of high-risk families.     

Local induction of angiotensin-converting enzyme in the kidney as a mechanism of progressive renal diseases

Angiotensin Converting Enzyme (ACE) or Kininase II has a pivotal role determining the local activity of the renin angiotensin and kallikrein kinin systems. Angiotensin II (Ang II), a main hormone of the renin system, has a well established participation as a renal injury agent in models of renal disease with tubulointerstitial fibrosis. Although, since its discovery, ACE has been known to convert Ang I to Ang II, and to inactivate bradykinin (BK), only recently has been emerged evidence for a role of BK with renal protective and antifibrotic effects opposing Ang II. Pertinent to the tubulointerstitial injury, where infiltration and proliferation of macrophages and fibroblast occur, ACE also regulates the levels of the natural hemoregulatory peptide, N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP). Owing the importance of tissue ACE, its distribution was studied in several models of renal injury. The results showed increased ACE in proximal tubules and ACE induction in the cell infiltrated tubulointerstitium (macrophages and myofibroblasts) of injured kidneys from hypokalemic, Goldblatt hypertensive, Ang II and phenylefrine infused rats, and in both human diabetic and membranous nephropathies. ACE, in addition to Ang II generation, may play a pathogenic role through the hydrolysis of BK and Ac-SDKP. Thus, local increase in ACE can be a novel mechanism involved in tubulointerstitial renal injury, providing, from an anatomical ground, a pathological basis for the putative deleterious effect of ACE in the diseased kidneys, and the beneficial effect of ACE inhibition.     

Heat shock protein expression in cerebral vessels after subarachnoid hemorrhage

OBJECTIVE: The mechanisms of cerebral vasospasm after subarachnoid hemorrhage (SAH) remain controversial. Recent data have implicated two small heat shock proteins (HSPs), namely HSP20 and HSP27, in the regulation of vascular tone. Increases in the phosphorylation of HSP20 are associated with vasorelaxation, and increases in the phosphorylation of HSP27 are associated with impaired vasorelaxation. Therefore, we hypothesized that alterations in the expression and/or phosphorylation of these two small HSPs might play a role in cerebral vasospasm after SAH. METHODS: A rat model of endovascular perforation was used to induce SAH. Middle cerebral arteries were harvested from control animals, sham-treated animals, and animals with SAH, 48 hours after SAH induction. Dose-response curves for endothelium-independent (sodium nitroprusside, 10(-8) to 10(-4) mol/L) and endothelium-dependent (bradykinin, 10(-10) to 10(-5) mol/L) relaxing agents were recorded ex vivo. Physiological responses were correlated with the expression and phosphorylation of HSP20 and HSP27 by using one- and two-dimensional immunoblots. RESULTS: There was impaired endothelium-independent and endothelium-dependent relaxation in cerebral vessels after SAH. These changes were associated with decreased expression of both total and phosphorylated HSP20 and increases in the amount of phosphorylated HSP27. CONCLUSION: In this model, impaired relaxation of cerebral vessels after SAH was associated with increases in the amount of phosphorylated HSP27 and decreases in the expression and phosphorylation of HSP20. These data are consistent with alterations in the expression and phosphorylation of these small HSPs in other models of vasospasm.     

The independent oncological role for cytoreductive nephrectomy in metastatic renal cell carcinoma: Prognostic features in the era of targeted therapies

Objectives

To describe the effects of cytoreductive nephrectomy (CN) on the natural course of metastatic renal cell carcinoma (mRCC). CN appears to stabilize metastatic lesions in mRCC in a subgroup of patients and we hypothesize that systemic treatment might be deferred in these patients with stable disease after CN.

Trends in internet search activity,media coverage,and patient-centered health information after the FDA safety communications on surgical mesh for pelvic organ prolapse

Introduction

In July 2011, the US Food and Drug Administration (FDA) issued a safety communication regarding serious complications associated with surgical mesh for pelvic organ prolapse, prompting increased media and public attention. This study sought to analyze internet search activity and news article volume after this FDA warning and to evaluate the quality of websites providing patient-centered information.

Methods

Google Trends? was utilized to evaluate search engine trends for the term “pelvic organ prolapse” and associated terms between 1 January 2004 and 31 December 2014. Google News? was utilized to quantify the number of news articles annually under the term “pelvic organ prolapse.” The search results for the term “pelvic organ prolapse” were assessed for quality using the Health On the Net Foundation (HON) certification.

Results

There was a significant increase in search activity from 37.42 in 2010 to 57.75 in 2011, at the time of the FDA communication (p?=?0.021). No other annual interval had a statistically significant increase in search activity. The single highest monthly search activity, given the value of 100, was August 2011, immediately following the July 2011 notification, with the next highest value being 98 in July 2011. Linear regression analysis of news articles per year since the FDA communication revealed r²?=?0.88, with a coefficient of 186. Quality assessment demonstrated that 42 % of websites were HON-certified, with .gov sites providing the highest quality information.

Conclusions

Although the 2011 FDA safety communication on surgical mesh was associated with increased public and media attention, the quality of relevant health information on the internet remains of poor quality. Future quality assurance measures may be critical in enabling patients to play active roles in their own healthcare.

     

Contemporary standards for the diagnosis and treatment of heparin-induced thrombocytopenia (HIT)

BACKGROUND: Heparin binding to platelet factor 4 (PF4) generates a new antigenic epitope. In an unpredictable fashion, as many as approximately 17% of patients treated with unfractionated heparin (UFH) and approximately 8% treated with low-molecular-weight heparin (LMWH) subsequently develop the anti-heparin-PF4 antibodies that mediate heparin-induced thrombocytopenia and thrombosis (HIT). Very few of those patients with circulating anti-heparin-PF4 antibodies, however, progress to develop clinical HIT (referred to previously as Type II HIT). Only 20% of those who harbor antibodies ( approximately 3% of those exposed to heparin) will manifest the thrombocytopenia subsequently. Even fewer patients (0.03% to 0.09% of those exposed to heparin) experience the marked platelet activation and morbid thromboses characteristic of the HIT syndrome. The pathogenesis of heparin-induced thrombocytopenia (HIT) remains elusive. The pathophysiologic understanding to date has revolved around pathogenic anti-heparin-PF4 antibodies that trigger platelet activation, release of platelet procoagulant microparticles, and resultant thrombosis. The clinical diagnosis of HIT is confusing because current assays to detect anti-heparin-PF4 antibodies do not correlate well with the disease. Currently available assays lack either adequate sensitivity and interlaboratory reproducibility (ie, functional serotonin release assays) or specificity (ie, enzyme-linked immunosorbent assays or ELISAs). CONCLUSIONS: Fortunately, the treatment for HIT is not confusing. The purposes of this review are as follows: (1) to examine the relevant clinical definition of HIT, (2) to explore our current understanding as to the pathogenesis of HIT, and (3) to present an algorithm for the identification and treatment of the HIT syndrome.     

The SF-36 health status questionnaire in assessing patients with epistaxis secondary to hereditary hemorrhagic telangiectasia

BACKGROUND: This study assesses the ability of the short form 36 (SF-36), a validated health status survey, to measure the health of patients with epistaxis due to hereditary hemorrhagic telangiectasia (HHT). METHODS: Thirty-eight patients completed the SF-36 and symptom-specific questionnaires. They were asked to rate their epistaxis as severe, moderate, or mild. The SF-36 data are compared with a reference population and analyzed with respect to the subgroups. RESULTS: The scores for each dimension of health were significantly reduced (p < 0.05) when compared with the reference population for all dimensions except pain. When compared with the patients' ratings of severity, a significant correlation was detected in five dimensions (p < 0.05). CONCLUSION: The SF-36 reflects the reduced health status of patients with epistaxis due to HHT. Changes in the SF-36 score could be used as an outcome measure in assessing efficacy of treatment of this condition.