Audit on the degree of application of universal precautions in a haemodialysis unit.

BACKGROUND: The purpose of the study was to investigate the degree of compliance with standard precautions (hand washing and wearing of gloves) by health workers in one haemodialysis unit. METHODS: During 4 months, two observers monitored the activities of the health care staff in the dialysis unit. Thirty five randomly distributed observation periods of 60 min duration covered one haemodialysis session. The observers evaluated (i) the total number of potential opportunities to implement standard precautions and (ii) the number of occasions when these were actually put into practice. RESULTS: A total of 364 opportunities to wear gloves and to wash hands thereafter and 273 opportunities to wash hands before a patient-oriented activity were observed. The proportion of occasions when gloves were actually used was 18.7%. Hand washing after a patient-oriented activity was performed only on 32.4% of occasions. Finally, only on 3% of such occasions was hand washing before the activity. CONCLUSIONS: The degree of compliance with standard precautions by health care personnel is unsatisfactory and this favours nosocomial transmission in haemodialysis units.     

Constitutive Phosphorylation as a Key Regulator of TRPM8 Channel Function

In mammals, environmental cold sensing conducted by peripheral cold thermoreceptor neurons mostly depends on TRPM8, an ion channel that has evolved to become the main molecular cold transducer. This TRP channel is activated by cold, cooling compounds, such as menthol, voltage, and rises in osmolality. TRPM8 function is regulated by kinase activity that phosphorylates the channel under resting conditions. However, which specific residues, how this post-translational modification modulates TRPM8 activity, and its influence on cold sensing are still poorly understood. By mass spectrometry, we identified four serine residues within the N-terminus (S26, S29, S541, and S542) constitutively phosphorylated in the mouse ortholog. TRPM8 function was examined by Ca²⁺ imaging and patch-clamp recordings, revealing that treatment with staurosporine, a kinase inhibitor, augmented its cold- and menthol-evoked responses. S29A mutation is sufficient to increase TRPM8 activity, suggesting that phosphorylation of this residue is a central molecular determinant of this negative regulation. Biophysical and total internal reflection fluorescence-based analysis revealed a dual mechanism in the potentiated responses of unphosphorylated TRPM8: a shift in the voltage activation curve toward more negative potentials and an increase in the number of active channels at the plasma membrane. Importantly, basal kinase activity negatively modulates TRPM8 function at cold thermoreceptors from male and female mice, an observation accounted for by mathematical modeling. Overall, our findings suggest that cold temperature detection could be rapidly and reversibly fine-tuned by controlling the TRPM8 basal phosphorylation state, a mechanism that acts as a dynamic molecular brake of this thermo-TRP channel function in primary sensory neurons.SIGNIFICANCE STATEMENT Post-translational modifications are one of the main molecular mechanisms involved in adjusting the sensitivity of sensory ion channels to changing environmental conditions. Here we show, for the first time, that constitutive phosphorylation of the well-conserved serine 29 within the N-terminal domain negatively modulates TRPM8 channel activity, reducing its activation by agonists and decreasing the number of active channels at the plasma membrane. Basal phosphorylation of TRPM8 acts as a key regulator of its function as the main cold-transduction channel, significantly contributing to the net response of primary sensory neurons to temperature reductions. This reversible and dynamic modulatory mechanism opens new opportunities to regulate TRPM8 function in pathologic conditions where this thermo-TRP channel plays a critical role.     

BRCA1/Trp53 heterozygosity and replication stress drive esophageal cancer development in a mouse model

BRCA1 germline mutations are associated with an increased risk of breast and ovarian cancer. Recent findings of others suggest that BRCA1 mutation carriers also bear an increased risk of esophageal and gastric cancer. Here, we employ a Brca1/Trp53 mouse model to show that unresolved replication stress (RS) in BRCA1 heterozygous cells drives esophageal tumorigenesis in a model of the human equivalent. This model employs 4-nitroquinoline-1-oxide (4NQO) as an RS-inducing agent. Upon drinking 4NQO-containing water, Brca1 heterozygous mice formed squamous cell carcinomas of the distal esophagus and forestomach at a much higher frequency and speed (∼90 to 120 d) than did wild-type (WT) mice, which remained largely tumor free. Their esophageal tissue, but not that of WT control mice, revealed evidence of overt RS as reflected by intracellular CHK1 phosphorylation and 53BP1 staining. These Brca1 mutant tumors also revealed higher genome mutation rates than those of control animals; the mutational signature SBS4, which is associated with tobacco-induced tumorigenesis; and a loss of Brca1 heterozygosity (LOH). This uniquely accelerated Brca1 tumor model is also relevant to human esophageal squamous cell carcinoma, an often lethal tumor.

Germline BRCA1 mutations predispose humans to an elevated cancer risk and especially that of the breast and ovary (1, 2). In recent times the suggestion of an increased risk of esophageal cancer in BRCA1 mutation carriers has also been reported, e.g., in an individual with a germline BRCA1 mutation (3). Also, a complete clinical response to platinum treatment was observed in a patient with BRCA1 mutant esophageal cancer (4). Furthermore, the overall esophageal squamous cell carcinoma (ESCC) risk in BRCA1 carriers is significant (relative risk [RR] of 2.9 [95% CI 1.1 to 6.0]) (5, 6). This is in keeping with the observation that a relatively frequent loss of heterozygosity (LOH) is detected in the BRCA1-containing region of chromosome 17 in squamous cell carcinoma of the esophagus (7–9).BRCA1 maintains genome integrity by engaging in multiple cellular processes, including the repair of DNA damage (10, 11), including double strand breaks (DSBs), stalled replication forks, and other abnormalities. Stalled forks, when not resolved, can lead to mutations or can collapse into DSBs (12–15). Both outcomes are components of what is commonly referred to as replication stress (RS), which, when chronic, can serve as a cancer driving force (16–18).Loss of certain DNA damage repair functions in BRCA1 mutant tumor cells also renders these cells sensitive to platinum-based derivatives and PARP inhibitors (19, 20). Success of these agents in suppressing BRCA1 mutant tumor growth has made them therapeutic agents of choice for treating BRCA1 mutant cancer (21, 22). Loss of BRCA1 function either by germline deletion and/or promoter hypermethylation is now a predictive classifier of response to these agents (23).Currently, multiple Brca1 mouse models facilitate the study of BRCA1 loss-associated tumorigenesis. Complete loss of BRCA1 is embryonically lethal (24); thus, successful, tumorigenic models either conditionally delete both alleles of Brca1 in a tissue of interest or express a hypomorphic mutant version of Brca1 (25–27). For example, conditional Brca1 loss can be driven by Cre-mediated deletion of two Brca1 floxed alleles in a tissue of choice. And mice bearing hypomorphic Brca1 mutant alleles, like delta 11, and those that express an incompletely functioning or a truncated version of the Brca1 protein instead of the full-length polypeptide also develop Brca1 tumors (26, 28, 29).Often, BRCA1 knockout (KO) mice also incur a loss of p53 function, which results in accelerated tumor formation, and, in the case of hypomorphic Brca1 mutant mice, their survival as well (25, 27, 30). Mice in both models develop tumors, including mammary and ovarian tumors, which, on average, take ∼1 y to develop. Brca1 delta 11 (Brca1^Δ11/Δ11) mice, which synthesize a markedly deleted but still partially functional Brca1 allele, also form esophageal tumors that can be accelerated by addition of the oxidative stress–inducing agent methyl-N-amylnitrosamine (MNAN) to their drinking water (31).However, the role of BRCA1 haploinsufficiency has not been extensively evaluated in mouse models. One reason is that Brca1 heterozygous mice did not generate tumors more often or faster than their wild-type (WT) counterparts (27, 32). This is unlike the increased predisposition to cancer observed in human BRCA1 mutation carriers (who carry a germline loss-of-function mutation in a single BRCA1 allele). They manifest a significantly greater than normal breast and/or ovarian cancer incidence by age 70 (1, 2).BRCA1 haploinsufficiency can be linked to increased genomic instability, in part, because of its defective participation in stalled fork repair and replication stress suppression (33) and, possibly, because of its role in regulating SIRT1 levels and affecting pRb pathway activation (34). Given the importance of RS development in tumorigenesis (16, 18, 35), this effect would be a logical contributor to BRCA1 mutant cancer development. Of note, BRCA1 heterozygous human cells are haploinsufficient for RS suppression (33), raising the possibility that this defect operates as a general contributor to the increased tumorigenicity observed in many germline BRCA1 heterozygous families.To test this hypothesis, we have established a Brca1 mutant esophageal mouse cancer model that is capable of addressing the role of replication stress accumulation in BRCA1 mutant cancer. Here one allele of Brca1 and one of Trp53 were deleted through the action of Meox2Cre, which acts very early during embryogenesis (embryonic day 5 [E5]) (36) and results in the development of Brca1 and p53 heterozygosity in all tissues. Using this mouse model, we have found that BRCA1 deficiency in replication stress suppression is enhanced by exposure to 4-nitroquinoline-1-oxide (4NQO) in BRCA1 heterozygous tissue where it serves as an efficient and abnormally rapid driver of tumor formation.     

Drug-mediated immunogenic changes of virus-induced leukemia in vivo.

LSTRA and RBL-5 lymphomas induced by Moloney and Rauscher leukemia viruses, respectively, were used to determine whether antigenically altered tumors induced by 5-(3,3-dimethyl-1-triazeno)-imidazole-4-carboxamide in vivo would retain their original antigenic properties and/or have new antigenic properties. The tumors became highly immunogenic in the syngeneic hosts after 4 to 8 transplant generations with drug treatment. Syngeneic mice could be protected against challenge with the parental tumor by presensitization with the drug-altered sublines while unrelated tumor lines were incapable of protecting them. The drug-altered subline of LSTRA was used for treatment of the LSTRA in conjunction with chemotherapy, and this immunochemotherapy produced significant increases in number of survivors and increases in median survival time compared to either treatment alone. Tolerance studies indicated that there are novel antigens and parental tumor antigens associated with the drug-treated sublines.     

A new interlocking dynamic compression nail for tibial shaft fractures

The Clos tibial nail is a cannulated cylindrical nail that permits static, dynamic as well as incompression mounting by the insertion of locking screws into distal and proximal holes. From September 1998 to March 1999 we treated 16 tibial shaft fractures with CLOS tibial nails. All fractures were managed with calcanear traction, closed reduction, reaming and fixation. Patients were followed for at least 1 year. The mean time to full weight bearing was 11 (10–14) weeks. There were no cases of delayed union or dynamization. All patients returned to their previous activity levels. Received: 4 September 2000; Accepted: 9 September 2000     

Adolescent physical performance and adult physical activity in Flemish males

Limited information is available about the associations between adolescent fitness levels and adult physical activity. In the present study, these associations are investigated using different indicators of physical activity. It is hypothesized that both health‐ and performance‐related fitness characteristics, observed during the adolescent period, contribute equally to the explained variance in adult physical activity levels. Subjects were 109 Flemish males followed over a period of 27 years from 13 to 40 years of age in the Leuven Longitudinal Study on Lifestyle Fitness and Health. Performance and health‐related fitness characteristics were observed during the growth period and at 40 years of age. The Work Index, Leisure Time Index, and Sport Index of the Baecke questionnaire were used as indicators of physical activity together with triaxial accelerometry. Multiple regression and discriminant analyses contrasting extreme quintiles of activity groupings were used to analyse the associations. Only the Baecke Sport Index showed consistent significant associations (R² = 0.03 to R² = 0.23) with adolescent fitness levels observed at 13, 15, and 18 years. When upper and lower quintiles were contrasted, fitness characteristics observed at the three age levels during adolescence were significantly different for each of the three indices of the Baecke questionnaire at 40 years of age. Lowest associations (R² = 0.09 to R² = 0.17) were found for the Work Index, followed by the Leisure Time Index (R² = 0.12 to R² = 0.28) and Sport Index (R² = 0.25 to R² = 0.43). Highest associations were evident for the 18‐ to 40‐year interval. Performance‐ and health‐related fitness characteristics explain equally well the variance in physical activity indicators. Am. J. Hum. Biol. 13:173–179, 2001. © 2001 Wiley‐Liss, Inc.     

Response to Marabotto et al.

Biopsies have important value in assessing for nonerosive reflux disease.