Management of Depression in Chronic Care Patients Using a Task-Sharing Approach in a Real-World Primary Health Care Setting in South Africa: Outcomes of a Cohort Study.

Depressive symptoms are common in South African primary care patients with chronic medical conditions, but are usually unrecognised and untreated. This study evaluated an integrated, task-sharing collaborative approach to management of depression comorbid with chronic diseases in primary health care (PHC) patients in a real-world setting. Existing HIV clinic counsellors provided a manualised depression counselling intervention with stepped-up referral pathways to PHC doctors for initiation of anti-depressant medication and/ or referral to specialist mental health services. Using a comparative group cohort design, adult PHC patients in 10 PHC facilities were screened with the Patient Health Questionnaire-9 with those scoring above the validated cut-off enrolled. PHC nurses independently assessed, diagnosed and referred patients. Referral for treatment was independently associated with substantial improvements in depression symptoms three months later. The study confirms the viability of task-shared stepped-up collaborative care for depression treatment using co-located counselling in underserved real-world PHC settings.

Brace yourselves: esports is coming.

Background: Competitive gaming (or esports) is an emerging phenomenon with a field of over 454 million fans globally. Despite its tremendous popularity and commercial support, esports is not widely understood. It is also disregarded as a reputable or credible form of competition. The International Olympic Committee (IOC) contends that esports may be considered a sporting activity, but this is limited to the basis of its sedentary nature and poor governance. Discussion: These authors present evidence to inform and clarify misconceptions surrounding esports among the broader scientific community. They also encourage researchers to engage in further work into the phenomenon of competitive gaming with regard to health and performance, resulting in a better understanding of esports and guiding its development as a credible, competitive entity.

A randomized trial and novel SPR technique identifies altered lipoprotein-LDL receptor binding as a mechanism underlying elevated LDL-cholesterol in APOE4s.

At a population level APOE4 carriers (~25% Caucasians) are at higher risk of cardiovascular diseases. The penetrance of genotype is however variable and influenced by dietary fat composition, with the APOE4 allele associated with greater LDL-cholesterol elevation in response to saturated fatty acids (SFA). The etiology of this greater responsiveness is unknown. Here a novel surface plasmon resonance technique (SPR) is developed and used, along with hepatocyte (with the liver being the main organ modulating lipoprotein metabolism and plasma lipid levels) uptake studies to establish the impact of dietary fatty acid composition on, lipoprotein-LDL receptor (LDLR) binding, and hepatocyte uptake, according to APOE genotype status. In men prospectively recruited according to APOE genotype (APOE3/3 common genotype, or APOE3/E4), triglyceride-rich lipoproteins (TRLs) were isolated at fasting and 4-6 h following test meals rich in SFA, unsaturated fat and SFA with fish oil. In APOE4s a greater LDLR binding affinity of postprandial TRL after SFA, and lower LDL binding and hepatocyte internalization, provide mechanisms for the greater LDL-cholesterol raising effect. The SPR technique developed may be used for the future study of the impact of genotype, and physiological and behavioral variables on lipoprotein metabolism. Trial registration number NCT01522482.

Sample introduction interface for on-chip nucleic acid-based analysis of Helicobacter pylori from stool samples.

Despite recent advances in microfluidic-based integrated diagnostic systems, the sample introduction interface, especially with regards to large volume samples, has often been neglected. We present a sample introduction interface that allows direct on-chip processing of crude stool samples for the detection of Helicobacter pylori (H. pylori). The principle of IFAST (immiscible filtration assisted by surface tension) was adapted to include a large volume sample chamber with a septum-based interface for stool sample introduction. Solid chaotropic salt and dry superparamagnetic particles (PMPs) could be stored on-chip and reconstituted upon sample addition, simplifying the process of release of DNA from H. pylori cells and its binding to the PMPs. Finally, the PMPs were pulled via a magnet through a washing chamber containing an immiscible oil solution and into an elution chamber where the DNA was released into aqueous media for subsequent analysis. The entire process required only 7 min while enabling a 40-fold reduction in working volume from crude biological samples. The combination of a real-world interface and rapid DNA extraction offers the potential for the methodology to be used in point-of-care (POC) devices.

Neuroprotective potential of pleiotrophin overexpression in the striatonigral pathway compared with overexpression in both the striatonigral and nigrostriatal pathways.

Intrastriatal injection of recombinant adeno-associated viral vector serotype 2/1 (rAAV2/1) to overexpress the neurotrophic factor pleiotrophin (PTN) provides neuroprotection for tyrosine hydroxylase immunoreactive (THir) neurons in the substantia nigra pars compacta (SNpc), increases THir neurite density in the striatum (ST) and reverses functional deficits in forepaw use following 6-hydroxydopamine (6-OHDA) toxic insult. Glial cell line-derived neurotrophic factor (GDNF) gene transfer studies suggest that optimal neuroprotection is dependent on the site of nigrostriatal overexpression. The present study was conducted to determine whether enhanced neuroprotection could be accomplished via simultaneous rAAV2/1 PTN injections into the ST and SN compared with ST injections alone. Rats were unilaterally injected in the ST alone or injected in both the ST and SN with rAAV2/1 expressing either PTN or control vector. Four weeks later, all rats received intrastriatal injections of 6-OHDA. Rats were euthanized 6 or 16 weeks relative to 6-OHDA injection. A novel selective total enumeration method to estimate nigral THir neuron survival was validated to maintain the accuracy of stereological assessment. Long-term nigrostriatal neuroprotection and functional benefits were only observed in rats in which rAAV2/1 PTN was injected into the ST alone. Results suggest that superior preservation of the nigrostriatal system is provided by PTN overexpression delivered to the ST and restricted to the ST and SN pars reticulata and is not improved with overexpression of PTN within SNpc neurons.

ARF inhibits the growth and malignant progression of non-small-cell lung carcinoma.

Non-small-cell lung carcinoma (NSCLC) is among the deadliest of human cancers. The CDKN2A locus, which houses the INK4a and ARF tumor suppressor genes, is frequently altered in NSCLC. However, the specific role of ARF in pulmonary tumorigenesis remains unclear. KRAS and other oncogenes induce the expression of ARF, thus stabilizing p53 activity and arresting cell proliferation. To address the role of ARF in Kras-driven NSCLC, we compared the susceptibility of NIH/Ola strain wild-type and Arf-knockout mice to urethane-induced lung carcinogenesis. Lung tumor size, malignancy and associated morbidity were significantly increased in Arf(-/-) compared with Arf(+/+) animals at 25 weeks after induction. Pulmonary tumors from Arf-knockout mice exhibited increased cell proliferation and DNA damage compared with wild-type mice. A subgroup of tumors in Arf(-/-) animals presented as dedifferentiated and metastatic, with many characteristics of pulmonary sarcomatoid carcinoma, a neoplasm previously undocumented in mouse models. Our finding of a role for ARF in NSCLC is consistent with the observation that benign adenomas from Arf(+/+) mice robustly expressed ARF, while ARF expression was markedly reduced in malignant adenocarcinomas. ARF expression also frequently colocalized with the expression of p21(CIP1), a transcriptional target of p53, arguing that ARF induces the p53 checkpoint to arrest cell proliferation in vivo. Taken together, these findings demonstrate that induction of ARF is an early response in lung tumorigenesis that mounts a strong barrier against tumor growth and malignant progression.

Relationship between the body composition and motor and physical competence of Grade 1 learners in South Africa.

AIM: The objective of this study was to determine relationships between body composition and motor and physical competence of Grade 1 learners living in the North West Province of South Africa. METHODS: Data were collected by means of a stratified random sampling procedure from 816 Grade 1 learners (419 boys, 397 girls) with a mean age of 6.84 years (+ 0.39 SD), in the NW-CHILD-study. Height, weight, skinfolds (subscapular, triceps, calf) and waist circumference were measured. International cut-off values were used to classify the subjects in normal, overweight or obese categories. Motor and physical competence was measured by using the Bruiniks-Oseretsky Test of Motor Proficiency-2 SF and the Test of Gross Motor Development 2. RESULTS: One out of 10 learners were overweight or obese. Fine motor precision, balance, running speed and agility and strength correlated significantly with BMI with no clear relationships with object control skills and upper limb-coordination. CONCLUSION: A clear relationship was found between body composition and most of the motor and physical fitness competencies of Grade 1 learners. Intervention strategies to improve the body composition of overweight children and accompanying motor proficiency back logs and physical fitness deficiencies are recommended.

The effects of Capn1 gene inactivation on skeletal muscle growth, development, and atrophy, and the compensatory role of other proteolytic systems.

Myofibrillar protein turnover is a key component of muscle growth and degeneration, requiring proteolytic enzymes to degrade the skeletal muscle proteins. The objective of this study was to investigate the role of the calpain proteolytic system in muscle growth development using µ-calpain knockout (KO) mice in comparison with control wild-type (WT) mice, and evaluate the subsequent effects of silencing this gene on other proteolytic systems. No differences in muscle development between genotypes were observed during the early stages of growth due to the up regulation of other proteolytic systems. The KO mice showed significantly greater m-calpain protein abundance (P < 0.01) and activity (P < 0.001), and greater caspase 3/7 activity (P < 0.05). At 30 wk of age, KO mice showed increased protein:DNA (P < 0.05) and RNA:DNA ratios (P < 0.01), greater protein content (P < 0.01) at the expense of lipid deposition (P < 0.05), and an increase in size and number of fast-twitch glycolytic muscle fibers (P < 0.05), suggesting that KO mice exhibit an increased capacity to accumulate and maintain protein in their skeletal muscle. Also, expression of proteins associated with muscle regeneration (neural cell adhesion molecule and myoD) were both reduced in the mature KO mice (P < 0.05 and P < 0.01, respectively), indicating less muscle regeneration and, therefore, less muscle damage. These findings indicate the concerted action of proteolytic systems to ensure muscle protein homeostasis in vivo. Furthermore, these data contribute to the existing evidence of the importance of the calpain system's involvement in muscle growth, development, and atrophy. Collectively, these data suggest that there are opportunities to target the calpain system to promote the growth and/or restoration of skeletal muscle mass.

Associations of genetic markers in cattle receiving differing implant protocols.

The potential interaction of growth-promoting implants and genetic markers previously reported to be associated with growth, carcass traits, and tenderness was evaluated. Two implant protocols were applied to subsets of steers (n = 383) and heifers (n = 65) that were also genotyped for 47 SNP reported to be associated with variation in growth, fat thickness, LM area, marbling, or tenderness. The "mild" protocol consisted of a single terminal implant [16 mg estradiol benzoate (EB), 80 mg trenbalone acetate (TBA) or 8 mg EB, 80 mg TBA given to steers and heifers, respectively]. The "aggressive" protocol consisted of both a growing implant (8 mg EB, 40 mg TBA) for the lightest half of the animals on the aggressive protocol and 2 successive implants (28 mg EB, 200 mg TBA) given to all animals assigned to the aggressive treatment. Implant protocol had measurable impact on BW and ADG (P < 0.05), with the aggressive protocol increasing these traits before the terminal implant (relative to the mild protocol), whereas the mild protocol increased ADG after the terminal implant so that the final BW and ADG over the experimental period were similar between protocols. Animals on the aggressive protocol had significantly increased (P < 0.05) LM area (1.9 cm(2)), slice shear force (1.4 kg), and intact desmin (0.05 units), but decreased (P < 0.05) marbling score (49 units) and adjusted fat thickness (0.1 cm), and yield grade (0.15 units). Among both treatments, 8 of 9 growth-related SNP were associated with BW or ADG, and 6 of 17 tenderness-related SNP were associated with slice shear force or intact desmin. Favorable growth alleles generally were associated with increased carcass yield traits but decreased tenderness. Similarly, favorable tenderness genotypes for some markers were associated with decreased BW and ADG. Some interactions of implant protocol and genotype were noted, with some growth SNP alleles increasing the effect of the aggressive protocol. In contrast, putative beneficial effects of favorable tenderness SNP alleles were mitigated by the effects of aggressive implant. These type of antagonisms of management variables and genotypes must be accounted for in marker assisted selection (MAS) programs, and our results suggest that MAS could be used to manage, but likely will not eliminate negative impact of implants on quality.

Effects of manipulation of the caspase system on myofibrillar protein degradation in vitro.

Apoptosis via the intrinsic caspase 9 pathway can be induced by oxidative stressors hydrogen peroxide (H2O2) and N-(4 hydroxyphenol) rentinamide (fenretinide), a synthetic retinoid. Accelerated muscle atrophy and proteolysis in muscle-wasting conditions have been linked to oxidative stress and activated protease systems. Therefore, the hypothesis of this study was that proteolysis of myofibrillar proteins could be manipulated through the induction or inhibition of the caspase system. After slaughter, LM and supraspinatus muscles from callipyge (n = 5) and normal (n = 3) lambs were excised, finely diced, and incubated with treatment buffers containing oxidative stressors fenretinide or H2O2, recombinant caspase 3, caspase-specific inhibitor N-acetyl-Asp-Glu-Val-Asp-CHO (DEVD), or control solution. Muscle samples were incubated for 1, 2, 7, and 21 d at 4°C. Activation of the initiator caspase, caspase 9, and myofibrillar protein degradation was determined by SDS-PAGE and Western blotting. Results showed that fenretinide, H2O2, and recombinant caspase 3 increased (P < 0.05) proteolysis of myofibril proteins, whereas DEVD inhibited degradation (P < 0.05). Proteolysis of myofibrillar proteins increased with incubation time (P < 0.0001), and incubation time × treatment interactions (P < 0.05) indicated that the treatment effects did not all occur at the same rate. This study has shown that manipulation of the caspase system through induction or inhibition of activity can affect degradation of myofibrillar proteins, providing further evidence that the caspase system could be involved in postmortem proteolysis and tenderization. However, these stimulated changes were not sufficient to overcome the lack of proteolysis that is characteristic of muscle from callipyge lambs.

Physical Activity Levels and Energy Expenditure of 9-Year-Old - 12-Year-Old Overweight and Obese Children

The high occurrence of overweight and obesity amongst children is a disturbing health problem worldwide. Possible causes of increasing childhood obesity are inactivity and energy imbalances. The aim of this study was to analyse the total energy expenditure (TEE) and physical activity levels in 9-year-old - 12-year-old overweight and obese children during a weekday and a weekend day; as well as during a weekday morning and afternoon. Twenty-four 9-year-old - 12-year-old children (seven boys and 17 girls); of whom nine were overweight and 15 were obese; were selected from seven public primary schools for this study. Body mass index (BMI) cut-off points were used to distinguish between overweight and obese. Each participant wore an ACTICALT monitor to determine their physical activity levels and TEE. It was found that the TEE of the children did not differ between a week day and a weekend day; although the TEE of the week day afternoon differed significantly from that of the weekday morning. Unlike the overweight children; none of the obese children met the requirements of 60 minutes of moderate-intensity physical activity per day. Strategies should be found to increase the activity levels of overweight and; especially; obese children; specifically during the mornings and over weekends

Contribution of genetic influences to animal-to-animal variation in myoglobin content and beef lean color stability.

Longissimus thoracis steaks from steers (n = 464) with 0 to 50% inheritance of Angus, Charolais, Gelbvieh, Hereford, Limousin, Red Angus, and Simmental were evaluated during 6 d of display to assess genetic contributions to color stability. Color space values [CIE L* (lightness), a* (redness), b* (yellowness)], chroma, color change (DeltaE), and surface metmyoglobin (K/S 572/525) were determined on d 0 and 6 of display. Myoglobin concentration was highly heritable (0.85), but ultimate pH was weakly heritable (0.06). Day 0 L* values were moderately heritable (0.24). Variation in metmyoglobin, L*, and DeltaE on d 6 was moderately explained by genetic factors (41, 40, and 29%, respectively). Change during display was moderately heritable for a* (0.31), b* (0.23), chroma (0.35), and surface metmyoglobin (0.29). At the start of display, Angus steaks had greater (P < 0.05) L* values than those from all breeds except Charolais. On d 6, Angus steaks had greater (P < 0.05) L* (50.0) values than Gelbvieh, Hereford, and Simmental steaks (46.1, 44.0, and 44.5, respectively). Day 0 values for a*, b*, chroma, and DeltaE were not affected by breed (P > 0.05). On d 6, a* values were greater (P < 0.05) for Charolais and Limousin steaks (31.1 and 30.5) than Angus, Hereford, and Red Angus steaks (27.4, 27.7, and 26.3, respectively). Thus, a* changed less (P < 0.05) in Charolais and Limousin steaks (1.8 and 2.6, respectively) vs. steaks from other breeds. Day 6 b* values were greater (P < 0.05) in Charolais (24.5) and Limousin steaks (24.0) vs. Gelbvieh (22.2), Hereford (21.9), and Red Angus steaks (21.4). Thus, b* values changed less (P < 0.05) in Charolais and Limousin steaks (1.5 and 1.7, respectively) than in Angus, Gelbvieh, Hereford, and Red Angus steaks (4.3, 3.8, 4.4, and 5.1, respectively). After 6 d of display, Charolais and Limousin steaks had greater chroma (P < 0.05; 39.5 and 38.8, respectively) compared with Angus, Hereford, and Red Angus steaks (35.4, 35.3, and 33.9, respectively). Less (P < 0.05) change in chroma occurred for Charolais and Limousin (2.1 and 2.8, respectively) than in Angus, Gelbvieh, Hereford, and Red Angus steaks (7.1, 6.6, 7.4, and 9.0, respectively). Myoglobin concentration was less for Charolais and Limousin (P < 0.05; 2.77 and 2.72, respectively) compared with Gelbvieh, Red Angus, and Simmental steaks (3.62, 3.43, and 3.71, respectively). Breeds did not differ in pH (P > 0.05). These data suggest Charolais- and Limousin-carcasses produced steaks with greater lean color stability than Angus, Hereford, and Red Angus carcasses. Furthermore, these findings suggest that genetics contribute substantially to animal-to-animal variation in lean color, particularly in maintaining color.

Inhibition of PI-3K restores nuclear p27Kip1 expression in a mouse model of Kras-driven lung cancer.

Reduced expression of the CDK inhibitor p27(Kip1) (p27) in human lung cancer correlates with tumor aggressiveness and poor prognosis. However, the regulation of p27 expression and the role of p27 during lung cancer are poorly understood. Urethane-induced lung tumors in mice frequently harbor mutations in the Kras oncogene, and in this study, we use this model to address the regulation of p27 during tumorigenesis. The Ras effector Akt is known to regulate p27 mRNA abundance by phosphorylating and inactivating the FOXO transcription factors. Phosphorylated Akt and FOXO proteins were both increased in lung tumors, correlating with a reduction in p27 mRNA transcript. Akt also directly phosphorylates p27 and regulates its nuclear/cytoplasmic localization. Tumors showed a reduced nuclear/cytoplasmic ratio of p27 protein, together with an increase in phosphorylated Thr197 p27 in the cytoplasmic pool. Treatment of lung tumor-bearing mice with the phosphoinositol-3 kinase inhibitor LY294002 induced a rapid decrease in phosphorylated Akt and phosphorylated p27, concomitant with an increase in nuclear p27. Germline p27 deficiency accelerated both the growth and malignant progression of urethane-induced lung tumors, and did so in a cell autonomous manner, confirming a causal role of p27 in tumor suppression. These results show that p27 is a potent barrier to the growth and malignant progression of Kras-initiated lung tumors. Further, the reduction of nuclear p27 in tumors is mediated by oncogene signaling pathways, which can be reversed by pharmacological agents.

Comparison of the pseudorabies virus Us9 protein with homologs from other veterinary and human alphaherpesviruses.

Pseudorabies virus (PRV) Us9 is a small, tail-anchored (TA) membrane protein that is essential for axonal sorting of viral structural proteins and is highly conserved among other members of the alphaherpesvirus subfamily. We cloned the Us9 homologs from two human pathogens, varicella-zoster virus (VZV) and herpes simplex virus type 1 (HSV-1), as well as two veterinary pathogens, equine herpesvirus type 1 (EHV-1) and bovine herpesvirus type 1 (BHV-1), and fused them to enhanced green fluorescent protein to examine their subcellular localization and membrane topology. Akin to PRV Us9, all of the Us9 homologs localized to the trans-Golgi network and had a type II membrane topology (typical of TA proteins). Furthermore, we examined whether any of the Us9 homologs could compensate for the loss of PRV Us9 in anterograde, neuron-to-cell spread of infection in a compartmented chamber system. EHV-1 and BHV-1 Us9 were able to fully compensate for the loss of PRV Us9, whereas VZV and HSV-1 Us9 proteins were unable to functionally replace PRV Us9 when they were expressed in a PRV background.

The caspase proteolytic system in callipyge and normal lambs in longissimus, semimembranosus, and infraspinatus muscles during postmortem storage.

The objective of this experiment was to determine whether the caspase proteolytic system has a role in postmortem tenderization. Six ewes and 6 wethers that were noncarriers and 6 ewes and 6 wethers that were expressing the callipyge gene were used for this study. Caspase activities were determined in LM at 7 different time points during the postmortem storage period: 0 h, 4 h, 8 h, 24 h, 2 d, 7 d, and 21 d and in semimembranosus (SM) and infraspinatus (IS) muscles at 0 h, 8 h, 24 h, and 7 d from callipyge and noncallipyge (normal) lambs. Calpastatin activity was determined at 0 h, 2 d, 7 d, and 21 d and slice shear force measured at 2, 7, and 21 d in the LM. Calpastatin activity and slice shear force were greater in LM from callipyge lambs than normal lambs at each time point (P < 0.001 and P < 0.0001, respectively). Caspases 3 and 7 are executioner caspases, and their combined activity was found to decrease during the postmortem storage period in LM, SM, and IS muscles from callipyge and normal lambs. Similarly, activity of the initiator caspase (caspase 9) decreased (P < 0.05) in all 3 muscles across the postmortem storage period in callipyge and normal lambs, and its decrease in activity preceded that of the executioner caspases 3/7. A positive relationship also was detected between caspase 9 and caspase 3/7 in LM, SM, and IS muscles (P < 0.0001, r = 0.85, r = 0.86, r = 0.84, respectively), which is consistent with caspase 9 being responsible for the cleavage and activation of the executioner caspases (caspase 3/7) downstream. Caspase 3/7 and caspase 9 activities at 8 h in SM were greater in normal lamb than callipyge lamb (P < 0.05), with a trend for caspase 3/7 activity to be greater at 24 h postmortem (P = 0.0841). There also was a trend for caspase 3/7 activity to be greater in LM at 21 d in normal lamb than in callipyge lamb (P = 0.053), although there were no differences detected in caspase activities between genotypes in the IS muscle, which is not affected by the callipyge gene. A negative relationship also was detected between peak caspase 3/7 activity at 8 h in LM from normal lambs and calpastatin activity at 0 and 2 d (r = -0.65, r = -0.68, respectively, P < 0.05). This relationship was not observed in LM from callipyge lambs, suggesting that caspase 3/7 may be cleaving calpastatin in normal lambs but the level of calpastatin in callipyge lambs is such that caspase 3/7 cannot degrade it sufficiently to overcome the increased content of calpastatin, and thus, calpastatin activity is the overriding factor in postmortem proteolysis in these animals. There was no direct evidence from this study that caspases have a significant role in postmortem tenderization, but they may have some role through calpastatin degradation.

Polymorphism in CYP17, GSTM1 and the progesterone receptor genes and its relationship with mammographic density

Radiologic breast density is one of the predictive factors for breast cancer and the extent of the density is directly related to postmenopause. However, some patients have dense breasts even during postmenopause. This condition may be explained by the genes that codify for the proteins involved in the biosynthesis, as well as the activity and metabolism of steroid hormones. They are polymorphic, which could explain the variations of individual hormones and, consequently, breast density. The constant need to find markers that may assist in the primary prevention of breast cancer as well as in selecting high risk patients motived this study. We determined the influence of genetic polymorphism of CYP17 (cytochrome P450c17, the gene involved in steroid hormone biosynthesis), GSTM1 (glutathione S-transferase M1, an enzyme involved in estrogen metabolism) and PROGINS (progesterone receptor), for association with high breast density. One hundred and twenty-three postmenopausal patients who were not on hormone therapy and had no clinical or mammographic breast alterations were included in the present study. The results of this study reveal that there was no association between dense breasts and CYP17 or GSTM1. There was a trend, which was not statistically significant (P = 0.084), towards the association between PROGINS polymorphism and dense breasts. However, multivariate logistic regression showed that wild-type PROGINS and mutated CYP17, taken together, resulted in a 4.87 times higher chance of having dense breasts (P = 0.030). In conclusion, in the present study, we were able to identify an association among polymorphisms, involved in estradiol biosyntheses as well as progesterone response, and radiological mammary density.

Polymorphism in CYP17, GSTM1 and the progesterone receptor genes and its relationship with mammographic density.

Radiologic breast density is one of the predictive factors for breast cancer and the extent of the density is directly related to postmenopause. However, some patients have dense breasts even during postmenopause. This condition may be explained by the genes that codify for the proteins involved in the biosynthesis, as well as the activity and metabolism of steroid hormones. They are polymorphic, which could explain the variations of individual hormones and, consequently, breast density. The constant need to find markers that may assist in the primary prevention of breast cancer as well as in selecting high risk patients motived this study. We determined the influence of genetic polymorphism of CYP17 (cytochrome P450c17, the gene involved in steroid hormone biosynthesis), GSTM1 (glutathione S-transferase M1, an enzyme involved in estrogen metabolism) and PROGINS (progesterone receptor), for association with high breast density. One hundred and twenty-three postmenopausal patients who were not on hormone therapy and had no clinical or mammographic breast alterations were included in the present study. The results of this study reveal that there was no association between dense breasts and CYP17 or GSTM1. There was a trend, which was not statistically significant (P = 0.084), towards the association between PROGINS polymorphism and dense breasts. However, multivariate logistic regression showed that wild-type PROGINS and mutated CYP17, taken together, resulted in a 4.87 times higher chance of having dense breasts (P = 0.030). In conclusion, in the present study, we were able to identify an association among polymorphisms, involved in estradiol biosyntheses as well as progesterone response, and radiological mammary density.

Insights into drug metabolism by cytochromes P450 from modelling studies of CYP2D6-drug interactions.

The cytochromes P450 (CYPs) comprise a vast superfamily of enzymes found in virtually all life forms. In mammals, xenobiotic metabolizing CYPs provide crucial protection from the effects of exposure to a wide variety of chemicals, including environmental toxins and therapeutic drugs. Ideally, the information on the possible metabolism by CYPs required during drug development would be obtained from crystal structures of all the CYPs of interest. For some years only crystal structures of distantly related bacterial CYPs were available and homology modelling techniques were used to bridge the gap and produce structural models of human CYPs, and thereby obtain useful functional information. A significant step forward in the reliability of these models came seven years ago with the first crystal structure of a mammalian CYP, rabbit CYP2C5, followed by the structures of six human enzymes, CYP1A2, CYP2A6, CYP2C8, CYP2C9, CYP2D6 and CYP3A4, and a second rabbit enzyme, CYP2B4. In this review we describe as a case study the evolution of a CYP2D6 model, leading to the validation of the model as an in silico tool for predicting binding and metabolism. This work has led directly to the successful design of CYP2D6 mutants with novel activity-including creating a testosterone hydroxylase, converting quinidine from inhibitor to substrate, creating a diclofenac hydroxylase and creating a dextromethorphan O-demethylase. Our modelling-derived hypothesis-driven integrated interdisciplinary studies have given key insight into the molecular determinants of CYP2D6 and other important drug metabolizing enzymes.

p19/Arf and p53 suppress sentinel lymph node lymphangiogenesis and carcinoma metastasis.

The ability of tumor cells to metastasize is increasingly viewed as an interaction between the primary tumor and host tissues. Deletion of the p19/Arf or p53 tumor suppressor genes accelerates malignant progression and metastatic spread of 7,12-dimethylbenz(a)anthracene (DMBA)/12-O-tetradecanoyl-phorbol-13-acetate (TPA)-induced squamous cell carcinomas, providing a model system to address mechanisms of metastasis. Here, we show that benign pre-metastatic papillomas from wild-type mice trigger lymphangiogenesis within draining lymph nodes, whereas there is no growth of primary tumor lymphatic vessels. Lymph node lymphangiogenesis is greatly accelerated in papilloma-bearing p19/Arf- or p53-deficient mice, which coincides with the greater propensity of these tumors to progress to carcinomas and to metastasize. The extent of accumulation of B cells within the tumor-draining lymph nodes of wild-type mice predicted the level of lymph node lymphangiogenesis and metastatic potential. Arf or p53 deficiency strongly accelerated lymph node immune cell accumulation, in a manner that was associated with the extent of lymph node lymphatic sinus growth. This immune cell accumulation and lymph node lymphangiogenesis phenotype identifies host anti-tumor responses that could drive metastatic spread of cancers via the lymphatics.