Remodeling of myocyte dimensions in hypertrophic and atrophic rat hearts.

Changes in hemodynamic load cause alterations in cardiac myocyte size, with regional variations in myocyte size distribution possible within the ventricular wall. We studied regional changes in cellular dimensions and their distribution in two models of cardiac hypertrophy and in cardiac atrophy in the rat. Combined volume-pressure overload was produced by 3,3',5-triiodo-L-thyronine (T3) treatment; atrophy was produced by heterotopic isotransplantation. Our previous data from long-term pressure overload after aortic constriction were used for comparison. Isolated ventricular myocytes were obtained after in vitro coronary perfusion with collagenase. Cell volume and its distribution were determined; cell length was directly measured by image analysis, and cross-sectional area was estimated from the cell volume/cell length ratio, assuming a cylindrical model. Myocyte hypertrophy resulting from hyperthyroidism and aortic constriction was primarily due to increased cross-sectional area. In both cases, the relative response was greater in the right ventricle than in the left ventricle. Within the left ventricle, epimyocardial myocytes enlarged the most. Aortic constriction and T3 treatment predominantly increased the size of smaller myocytes. Heterogeneity in myocyte size increased after constriction but remained relatively unaffected after T3 treatment. Atrophy of left ventricular myocytes was due to a proportional decrease in cell length and cross-sectional area, with the greatest decrease in the left ventricular endomyocardium. Atrophy predominantly affected larger myocytes, resulting in a more homogeneous overall population of smaller myocytes. We conclude that various alterations in load lead to diverse remodeling in the myocyte population throughout the ventricular wall. In general, smaller myocytes show the highest growth potential, whereas larger myocytes exhibit the highest potential to atrophy.     

Compound heterozygosity for unstable hemoglobin Genova and beta(o)-thalassemia associated with early onset of thalassemia major syndrome

We described the case of an infant with compound heterozygozity for a b0-thalassemic mutation and Hemoglobin (Hb) Genova, an unstable Hb variant. He has required regular transfusions as early as the second month of life and since then, behaves like a thalassemia major patient. This association leads to the most severe clinical course involving an unstable variant, reported so far.     

Comparison of the acute response to meals enriched with cis- or trans-fatty acids on glucose and lipids in overweight individuals with differing FABP2 genotypes

Trans-fatty acids have been implicated as a risk factor for cardiovascular disease and diabetes. In addition, a polymorphism at codon 54 (Ala54Thr) in the fatty acid-binding protein 2 (FABP2) gene has been suggested to modify an interaction between dietary fat and insulin sensitivity. We examined the postprandial metabolic profiles after meals enriched with C18:1trans- relative to a similar meal with C18:1cis-fatty acid in individuals who were either FABP2 Ala54 homozygotes or Thr54 carriers. Moderately overweight men and women ate 2 breakfast test meals, separated by 1 week, each providing 40% of their daily energy requirement and containing 50% of energy as fat. In one meal, 10% of energy was from C18:1trans, and in the other meal, the C18:1trans was replaced with C18:1cis. Metabolic parameters were assessed during an 8-hour period. Insulin and C-peptide levels increased more after the C18:1trans meal, and this was associated with a greater fall in free fatty acids. Postprandial glucose levels and oxidation of fatty acids and carbohydrate were not different between the 2 test meals. The Thr54 allele for FABP2 increased the rise in postprandial glucose but not triacylglycerols. Fractional triacylglycerol synthetic rates were higher after consumption of the C18:1trans meal relative to the C18:1cis meal only in Thr54 carriers. These data show that a single meal enriched with C18:1trans-fatty acids can significantly increase insulin resistance, and that in the presence of the FABP2 Thr54 allele, may contribute to increased partitioning of glucose to triacylglycerols and insulin resistance.     

Structural basis for p50RhoGAP BCH domain–mediated regulation of Rho inactivation

Spatiotemporal regulation of signaling cascades is crucial for various biological pathways, under the control of a range of scaffolding proteins. The BNIP-2 and Cdc42GAP Homology (BCH) domain is a highly conserved module that targets small GTPases and their regulators. Proteins bearing BCH domains are key for driving cell elongation, retraction, membrane protrusion, and other aspects of active morphogenesis during cell migration, myoblast differentiation, and neuritogenesis. We previously showed that the BCH domain of p50RhoGAP (ARHGAP1) sequesters RhoA from inactivation by its adjacent GAP domain; however, the underlying molecular mechanism for RhoA inactivation by p50RhoGAP remains unknown. Here, we report the crystal structure of the BCH domain of p50RhoGAP Schizosaccharomyces pombe and model the human p50RhoGAP BCH domain to understand its regulatory function using in vitro and cell line studies. We show that the BCH domain adopts an intertwined dimeric structure with asymmetric monomers and harbors a unique RhoA-binding loop and a lipid-binding pocket that anchors prenylated RhoA. Interestingly, the β5-strand of the BCH domain is involved in an intermolecular β-sheet, which is crucial for inhibition of the adjacent GAP domain. A destabilizing mutation in the β5-strand triggers the release of the GAP domain from autoinhibition. This renders p50RhoGAP active, thereby leading to RhoA inactivation and increased self-association of p50RhoGAP molecules via their BCH domains. Our results offer key insight into the concerted spatiotemporal regulation of Rho activity by BCH domain–containing proteins.

Small GTPases are molecular switches that cycle between an active GTP-bound state and an inactive GDP-bound state and are primarily involved in cytoskeletal reorganization during cell motility, morphogenesis, and cytokinesis (1, 2). These small GTPases are tightly controlled by activators and inactivators, such as guanine nucleotide exchange factors (GEFs) and GTPase-activating proteins (GAPs), respectively (3, 4), which are multidomain proteins that are themselves regulated through their interactions with other proteins, lipids, secondary messengers, and/or by posttranslational modifications (5–7). Despite our understanding of the mechanisms of action of GTPases, GAPs, and GEFs, little is known about how they are further regulated by other cellular proteins in tightly controlled local environments.The BNIP-2 and Cdc42GAP Homology (BCH) domain has emerged as a highly conserved and versatile scaffold protein domain that targets small GTPases, their GEFs, and GAPs to carry out various cellular processes in a spatial, temporal, and kinetic manner (8–15). BCH domain–containing proteins are classified into a distinct functional subclass of the CRAL_TRIO/Sec14 superfamily, with ∼175 BCH domain–containing proteins (in which 14 of them are in human) present across a range of eukaryotic species (16). Some well-studied BCH domain–containing proteins include BNIP-2, BNIP-H (CAYTAXIN), BNIP-XL, BNIP-Sα, p50RhoGAP (ARHGAP1), and BPGAP1 (ARHGAP8), with evidence to show their involvement in cell elongation, retraction, membrane protrusion, and other aspects of active morphogenesis during cell migration, growth activation and suppression, myoblast differentiation, and neuritogenesis (17–21). Aside from interacting with small GTPases and their regulators, some of these proteins can also associate with other signaling proteins, such as fibroblast growth factor receptor tyrosine kinases, myogenic Cdo receptor, p38-MAP kinase, Mek2/MP1, and metabolic enzymes, such as glutaminase and ATP-citrate lyase (17–26). Despite the functional diversity and versatility of BCH domain–containing proteins, the structure of the BCH domain and its various modes of interaction remain unknown. The BCH domain resembles the Sec14 domain (from the CRAL-TRIO family) (16, 27, 28), a domain with lipid-binding characteristics, which may suggest that the BCH domain could have a similar binding strategy. However, to date, the binding and the role of lipids in BCH domain function remain inconclusive.Of the BCH domain–containing proteins, we have focused on the structure and function of p50RhoGAP. p50RhoGAP comprises an N-terminal BCH domain and a C-terminal GAP domain separated by a proline-rich region. We found that p50RhoGAP contains a noncanonical RhoA-binding motif in its BCH domain and is associated with GAP-mediated cell rounding (13). Further, we showed previously that deletion of the BCH domain dramatically enhanced the activity of the adjacent GAP domain (13); however, the full dynamics of this interaction is unclear. Previously, it has been reported that the BCH and other domains regulate GAP activity in an autoinhibited manner (18, 21, 29, 30) involving the interactions of both the BCH and GAP domains, albeit the mechanism remains to be investigated. It has also been shown that a lipid moiety on Rac1 (a Rho GTPase) is necessary for its inactivation by p50RhoGAP (29, 31), which may imply a role in lipid binding. An understanding of how the BCH domain coordinates with the GAP domain to affect the local activity of RhoA and other GTPases would offer a previously unknown insight into the multifaceted regulation of Rho GTPase inactivation.To understand the BCH domain–mediated regulation of p50RhoGAP and RhoA activities, we have determined the crystal structure of a homologous p50RhoGAP BCH domain from S. pombe for functional interrogation. We show that the BCH domain adopts an intertwined dimeric structure with asymmetric monomers and harbors a unique RhoA-interacting loop and a lipid-binding pocket. Our results show that the lipid-binding region of the BCH domain helps to anchor the prenylation tail of RhoA while the loop interacts directly with RhoA. Moreover, we show that a mutation in the β5-strand releases the autoinhibition of the GAP domain by the BCH domain. This renders the GAP domain active, leading to RhoA inactivation and the associated phenotypic effects in yeast and HeLa cells. The released BCH domain also contributes to enhanced p50RhoGAP–p50RhoGAP interaction. Our findings offer crucial insights into the regulation of Rho signaling by BCH domain–containing proteins.     

Differences in the management and prognosis of women and men who suffer from acute coronary syndromes.

OBJECTIVES: The purpose of this research was to determine if sex and gender differences in the management of acute coronary syndromes (ACS) are associated with differences in prognosis after ACS. BACKGROUND: Previous investigators have reported sex/gender differences in the management of patients with ACS, but the impact of these differences on prognosis is unclear. METHODS: We analyzed data from the Clopidogrel in Unstable Angina to Prevent Recurrent Events (CURE) trial, which enrolled 4,836 women and 7,726 men with ACS. Patients were classified into risk strata using the Thrombolysis In Myocardial Infarction (TIMI) score. RESULTS: Women underwent fewer invasive procedures including angiography, angioplasty, and coronary artery bypass graft (CABG) surgery (47.6% vs. 60.5%; p = 0.0001) compared to men. No significant differences in cardiovascular death, myocardial infarction (MI), or stroke were observed (9.8% vs. 10.9%; p = 0.04), although women were more likely than men to develop refractory ischemia and to be rehospitalized for chest pain during follow-up (16.6% vs. 13.9%; p = 0.0001). These differences were particularly evident among TIMI high-risk women. A significant interaction between TIMI risk and gender for the outcome of refractory angina and rehospitalization for angina was present. CONCLUSIONS: Compared to men, high-risk women with ACS undergo less coronary angiography, angioplasty, and CABG surgery, and while they do not have higher incidence cardiovascular death, recurrent MI, or stroke, they suffer an increased rate of refractory ischemia and rehospitalization. All high-risk women and men with ACS should receive optimal medical management, and be considered for coronary angiography with possible revascularization if their coronary anatomy warrants it.     

The plaque lipid lysophosphatidic acid stimulates platelet activation and platelet-monocyte aggregate formation in whole blood: involvement of P2Y1 and P2Y12 receptors

Despite the fact that lysophosphatidic acid (LPA) has been identified as a main platelet-activating lipid of mildly oxidized low-density lipoprotein (LDL) and human atherosclerotic lesions, it remains unknown whether it is capable of activating platelets in blood. We found that LPA at concentrations slightly above plasma levels induces platelet shape change, aggregation, and platelet-monocyte aggregate formation in blood. 1-alkyl-LPA (16:0 fatty acid) was almost 20-fold more potent than 1-acyl-LPA (16:0). LPA directly induced platelet shape change in blood and platelet-rich plasma obtained from all blood donors. However, LPA-stimulated platelet aggregation in blood was donor dependent. It could be completely blocked by apyrase and antagonists of the platelet adenosine diphosphate (ADP) receptors P2Y1 and P2Y12. These substances also inhibited LPA-induced aggregation of platelet-rich plasma and aggregation and serotonin secretion of washed platelets. These results indicate a central role for ADP-mediated P2Y1 and P2Y12 receptor activation in supporting LPA-induced platelet aggregation. Platelet aggregation and platelet-monocyte aggregate formation stimulated by LPA was insensitive to inhibition by aspirin. We conclude that LPA at concentrations approaching those found in vivo can induce platelet shape change, aggregation, and platelet-monocyte aggregate formation in whole blood and suggest that antagonists of platelet P2Y1 and P2Y12 receptors might be useful preventing LPA-elicited thrombus formation in patients with cardiovascular diseases.     

Value of preoperative investigations in diagnosing prosthetic joint infection: retrospective cohort study and literature review

The diagnosis of a prosthetic joint infection is difficult, but crucial for appropriate treatment. Scintigraphy with specific markers for infection (labelled white cells or immunoglobulin-G) has been reported as a more reliable diagnostic tool than clinical assessment (fever, fistula), laboratory studies (polynuclear neutrophil count, erythrocyte rate sedimentation, and C-reactive protein), and preoperative aspiration. In the first part of this study, we retrospectively reviewed 230 patients admitted with a suspected prosthetic joint infection, and examined the validity of the different diagnostic tools for the group as a whole and for subgroups according to the Coventry classification. In the second part, we reviewed 35 articles about preoperative evaluation of infection in prosthetic joints and compared them to our findings. Our study indicates that C-reactive protein and joint aspiration are the most useful tools to diagnose prosthetic joint infection even in situations of chronic infection (Coventry type II).     

Reappraisal of the effectiveness of methotrexate in psoriatic arthritis: results from a longitudinal observational cohort

OBJECTIVE: In a previous study in our clinic, methotrexate (MTX) conferred no advantage with respect to clinical response or progression of damage after 24 months in patients with psoriatic arthritis (PsA). Our aim was to determine if MTX is being used earlier in the course of PsA and in a higher dose and whether that has led to improved outcomes. METHODS: All patients treated with MTX for at least 24 months in our clinic, between 1994 and 2004, were included in the study. The outcome measures were the progression of radiographic peripheral joint damage score and a > or = 40% reduction in the number of actively inflamed joints. The data from our study were compared to those obtained from our previous study. RESULTS: Fifty-nine patients (36 men) treated with MTX for 24 months were identified. The mean age was 46 years, PsA duration 8 years, and active joint count 12.1 (4.6 swollen). The mean increase in radiographic damage score was 1.5. Sixty-eight percent of patients demonstrated improvement at 24 months. When compared to our previous study, there was a trend for MTX to be used earlier, at a higher dose, with greater clinical improvement and less progression of damage. CONCLUSION: Our study suggests that treatment with MTX has changed in the past decade to include patients with shorter disease duration and less damage, at increased dose, and that there may be better response with less progression of damage.