Rare cardiomyopathies: diagnostic features.

Cardiomyopathies (CM) are an important and heterogeneous group of diseases affecting the myocardium. They can induce mechanical and/or electrical disorders and are due to a variety of causes, they frequently are genetic. However, since their high number and their clinical complexity, the identification is still a challenge. Echocardiography is a very useful tool in the assessment of CM. In this review we aim to define the typical clinical features and to discuss the main diagnostic tool, above all echocardiography that can help physicians in the correct assessment of CM.

Molecular basis of von Willebrand disease type IIB. Candidate mutations cluster in one disulfide loop between proposed platelet glycoprotein Ib binding sequences.

Many variants of von Willebrand disease (vWD) with qualitatively abnormal von Willebrand factor (vWF) are recognized. In vWD type IIB, the abnormal protein displays enhanced affinity for a platelet vWF receptor, the glycoprotein Ib-IX complex. 14 patients from 7 unrelated families with vWD type IIB were studied to determine the molecular basis for this phenotype. Specific oligonucleotide primers were used to amplify portions of vWF exon 28 encoding a domain that interacts with the platelet glycoprotein Ib-IX complex. Candidate missense mutations were identified for all 14 patients by DNA sequencing, allele specific oligonucleotide hybridization, and restriction endonuclease digestion. These sequence changes occur in an 11 amino acid segment within a single disulfide loop bounded by Cys(509) and Cys(695). All of these sequence changes are C----T transitions within CG dinucleotides. Six patients from two unrelated families were heterozygous for the encoded sequence Arg(543)----Trp. Seven patients from four unrelated families were heterozygous for the encoded sequence Arg(545)----Cys; this sequence change appears to have occurred independently three times, once as a new spontaneous mutation. One patient with apparently sporadic vWD type IIB was heterozygous for the encoded sequence Val(553)----Met, and this appears to be a new mutation. None of these sequence changes was found in 100 normal alleles. These findings suggest that vWD type IIB may be caused by relatively few distinct mutations, that these mutations may cluster within a specific region of one disulfide loop in vWF domain A1, and that this region can modulate the affinity of vWF for the platelet glycoprotein Ib-IX complex.

Alprazolam as a neuroleptic adjunct in the emergency treatment of schizophrenia.

OBJECTIVE: While neuroleptics remain the mainstay of drug intervention in the emergency management of psychosis, a variety of agents have received study as alternatives or adjuncts to these drugs in an attempt to improve the safety and efficacy of acute treatment. The purposes of this study were to investigate the efficacy and safety of alprazolam as a neuroleptic adjunct for schizophrenic patients in psychotic relapse and to clarify the effects of combination treatment on specific aspects of the psychotic process. METHOD: Twenty-eight acutely psychotic patients with schizophrenia who were admitted to an emergency psychiatric service were randomly assigned to treatment with either haloperidol and alprazolam or haloperidol with placebo under double-blind conditions. Drug administration lasted 72 hours. RESULTS: Both groups improved significantly. The combination-treated group required significantly less medication and had 56% fewer dystonic reactions. The addition of alprazolam was most effective for symptoms of excitement and uncooperativeness, particularly in the initial hours of treatment. CONCLUSIONS: The combination of alprazolam and haloperidol seems to be the most effective for agitated patients, particularly in the first 48 hours of treatment. It may also result in fewer dystonic reactions.

Characterization of partial gene deletions in type III von Willebrand disease with alloantibody inhibitors.

von Willebrand factor gene deletions were characterized in four patients with severe type III von Wilebrand disease and alloantibodies to von Willebrand factor. A PCR-based strategy was used to characterize the boundaries of the deletions. Identical 30 kb von Willebrand factor gene deletions which include exons 33 through 38 were identified in two siblings of one family by this method. A small 5 base pair insertion (CCTGG) was sequenced at the deletion breakpoint. PCR analysis was used to detect the deletion in three generations of the family, including two family members who are heterozygous for the deletion. In a second family, two type III vWD patients, who are distant cousins, share an approximately 56 kb deletion of exons 22 through 43. The identification and characterization of large vWF gene deletions in these type III vWD patients provides further support for the association between large deletions in both von Willebrand factor alleles and the development of inhibitory alloantibodies.

Effects of Prior Physical Activity on Skin Surface Temperature Response of the Ankle During and After a 30-minute Ice Pack Application.

The effects of various durations of treadmill running on ankle skin temperature response during and following ice application were investigated. We measured the ankle skin surface temperature of 12 male subjects with a telethermister (YSI Model 44) during each of three conditions: control and two exercise conditions that involved treadmill running for 15 or 30 minutes, followed by a 30-minute ice pack application and 90 minutes rewarming. The control condition involved no exercise prior to ice pack application. Ankle skin temperature increased significantly during 15 and 30 minutes of exercise, although the temperature difference between the two conditions was not statistically significant. Mean skin cooling temperatures were slightly, though not significantly, higher following exercise than following no exercise. The rate of cooling, however, was unaffected by prior exercise. Mean skin temperatures during rewarming were significantly higher following the exercise conditions, but the rate of rewarming was unchanged by exercise. Mean rewarming temperatures were higher in the 30-minute exercise condition than in the 15-minute exercise condition. Longer ice applications or shorter reapplications may be necessary following exercise of at least 15 minutes, but further investigation is necessary to substantiate this supposition.

Delirium.

Delirium is a heterogeneous syndrome that has multiple etiologies and is frequently underdiagnosed. This paper describes the varied symptoms, work-up, and management of delirium.

Impact of continuous value of body mass index on graft loss in overweight patients.

INTRODUCTION: High body mass index (BMI) is associated with increased cardiovascular mortality and risk of progression to end-stage renal disease both among the general population and among renal transplant patients. However, in the latter condition no unequivocal studies have been reported in the literature. The aim of our study was to investigate continuous versus categorical values of BMI (World Health Organization classification) as an independent risk factor in renal transplantation. PATIENTS AND METHODS: We retrospectively studied 194 renal transplant patients (128 males and 66 females) whose mean age at transplant was 43.9 years. They had 5 years follow-up. To investigate the association between BMI and graft survival, we performed univariate and multivariate analyses using the Cox regression model. This model was adjusted both for classical covariates (age, gender, time on dialysis, HLA mismatches, donor status) and other covariates as delayed graft function (DGF), acute rejection episodes (AR), and chronic allograft nephropathy (CAN), which are universally recognized to be predictors of graft loss as evidenced by a need for dialysis treatments. RESULTS: At the time of transplantation, the BMI averaged 24.4 +/- 2.65 kg/m(2). Upon univariate analysis, age (P = .049), BMI (P = .005), DGF (P = .009), ARE (P < .0001), and CAN (P = .001) were significantly related to poor transplant outcomes. Upon multivariate analysis, only the BMI value, considered as continuous value (P = .013), DGF (P = .030), and ARE (P < .0001) were significantly related to graft loss. CONCLUSIONS: BMI as a continuous value represented an independent risk factor for renal transplant loss at 5 years. Correction of pretransplant body weight both in overweight (25

Decisional trees in renal transplant follow-up.

INTRODUCTION: The predictive potentialities of application of data mining algorithms to medical research are well known. In this article, we have applied to a transplant population classification trees to build predictive models of graft failure, evaluating the interactions between body mass index (BMI) and other risk factors. The decision trees have been widely used to represent classification rules in a population by a hierarchical sequential structure. PATIENTS AND METHODS: We retrospectively studied 194 renal transplant patients with 5 years of follow-up (128 males, 66 females, mean age at time of transplant of 43.9 +/- 12.5 years). Exclusion criteria were: age < 18 years, multiorgan transplant, and retransplant. The BMI was calculated at the time of transplantation. In the classification algorithm, we considered the following parameters: age, sex, time on dialysis, donor type, donor age, HLA mismatches, delayed graft function (DGF), acute rejection episode (ARE), and chronic allograft nephropathy (CAN). The primary endpoint was graft loss within 5-years follow-up. RESULTS: The classification algorithm produced a decision tree that allowed us to evaluate the interactions between ARE, DGF, CAN, and BMI on graft outcomes, producing a validation set with 88.2% sensitivity and 73.8% specificity. Our model was able to highlight that subjects at risk of graft loss experienced one or more events of ARE, developed DGF and CAN, or has a BMI > 24.8 kg/m(2) and CAN. CONCLUSIONS: The use of decision trees in clinical practice may be a suitable alternative to the traditional statistical methods, since it may allow one to analyze interactions between various risk factors beyond the previous knowledge.

Prediction of chronic allograft nephropathy using classification trees.

INTRODUCTION: For its intrinsic potential to mine causal relations, machine learning techniques are useful to identify new risk indicators. In this work, we have shown two classification trees to predict chronic allograft nephropathy (CAN), through an evaluation of routine blood and urine tests. METHODS: We retrospectively analyzed 80 renal transplant patients with 60-month follow-up (mean = 55.20 +/- 12.74) including 52 males and 28 females of overall average age of 41.65 +/- 12.52 years. The primary endpoint was biopsy-proven CAN within 5 years from transplantation (n = 16). Exclusion criteria were multiorgan transplantations, patients aged less than 18 years, graft failure, or patient death in the first 6 months posttransplantation. Classification trees based on the C 4.8 algorithm were used to predict CAN development starting from patient features at transplantation and biochemical test at 6-month follow-up. Model performance was showed as sensitivity (S), false-positive rate (FPR), and area under the receiver operating characteristic curve (AUC). RESULTS: The two class of patients (no CAN versus CAN) showed significant differences in serum creatinine, estimated Glomerular Filtration Rate with Modification of Diet in Renal Disease study formula (MDRD), serum hemoglobin, hematocrit, blood urea nitrogen, and 24-hour urine protein excretion. Among the 23 evaluated variables, the first model selected six predictors of CAN, showing S = 62.5%, TFP = 7.2%, and AUC = 0.847 (confidence interval [CI] 0.749-0.945). The second model selected four variables, showing S = 81.3%, TFP = 25%, and AUC = 0.824 (CI 0.713-0.934). CONCLUSIONS: Identification models have predicted the onset of multifactorial, complex pathology, like CAN. The use of classification trees represent a valid alternative to traditional statistical models, especially for the evaluation of interactions of risk factors.

Kidney transplantation at Annunziata Hospital of Cosenza: report of 10 years experience.

OBJECTIVE: Kidney transplantation represents the gold standard for treatment of patients with end-stage renal disease. Herein we sought to report our 10-year experience with cadaveric kidney transplantations. PATIENTS AND METHODS: From February 1995 to September 2008, we performed 115 kidney transplantations. Patients were followed for an average of 4.9 years (range, 2.2-10.6 years). The cold ischemia time (CIT) averaged 13 +/- 3 hours, while the mean warm ischemic time was 25 +/- 10 minutes. The ureteral-bladder anastomosis was performed using Bracci catheters in the first series of 72 transplants, and double-J stents in the other 41 cases. The average waiting time was 122 +/- 21 months. The immunological regimens were prescribed according to the American Society of Nephrology (K/DOQI) with reference to comorbidity and concomitant risk factors and reported drug toxicity events. We transplanted kidneys with anatomic variations, ie, multiple arteries and double veins, and one double transplant of marginal organs. RESULTS: Our overall complication rate was 9.18%. The 10-year patient and graft survival rates were 89% and 84%, respectively. The percentage of biopsy-proven acute rejection episodes was 22.16%, while chronic allograft nephropathy (CAN) accounted for 15.3% at 5 years. The incidence of delayed graft function (DGF) was 14.05%. Finally, we noted 3 cases of cardiovascular death. CONCLUSION: Our experience showed excellent patient outcomes compared with other Italian and European data.

Angiotensin antagonists and fish oil for treating IgA nephropathy.

In IgA nephropathy (IgAN), ACE inhibitors (ACE-I) and angiotensin receptor blockers (ARB) are beneficial against hypertension, and their anti-proteinuric effect has been clearly demonstrated. However, sub-analyses of IgAN patients enrolled in large studies failed to prove a benefit against progression to renal failure. The European Community Biomed Concerted Action - a placebo-controlled randomized controlled trial begun in 1995 - in children and adults (9-35 years old) with proteinuria > 1 < 3.5 g/day/1.73 m(2) and normal or moderately reduced renal function proved the significant benefit of ACE-I on progression of kidney disease. The combination of ACE-I and ARB in proteinuric normotensive IgAN patients showed greater antiproteinuric effect and the COOPERATE trial also reported a superior effect of combination therapy in protecting against renal function deterioration. Treating IgAN with fish oil has a good rationale for renal inflammation as well as for prevention of cardiovascular morbidity. However, the published reports gave conflicting conclusions and also very recent data did not show significant benefits. In conclusion, ACE-I and ARB have a definite role in treating IgAN, particularly the hypertensive and proteinuric forms. These patients should be treated to target BP to <130/70 mm Hg and proteinuria <0.5 g/day.

Characterization of the Na+/H+ exchanger in human epidermoid carcinoma A431 vesicles.

A previous report from this laboratory (Rothenberg et al., 1983a) demonstrated the presence of an Na+/H+ exchanger in human epidermoid carcinoma A431 cells. We now characterize surface-derived membrane vesicles from this cell line which contain a functional Na+/H+ exchanger. The Na+/H+ exchanger in A431 vesicles shares a number of characteristics in common with previously described Na+/H+ exchangers including the following: (1) Na+ uptake is stimulated by an outward-directed pH gradient and inhibited by an inward-directed pH gradient. (2) Na+ uptake is inhibited by amiloride and its analogs and their relative effectiveness is similar in vesicles and A431 cells. (3) The Na+/H+ exchanger uses Na+ or Li+ as a substrate but not K+ or Cs+. (4) H+ efflux is stimulated by an inward-directed Na+ gradient and inhibited by the amiloride analog 5-N-dimethylamiloride. The Na+/H+ exchanger in these membrane vesicles is activated allosterically by low intravesicular pH. The apparent pKa of the activating site is 6.4-6.6, characteristic of the NA+/H+ exchanger before activation by mitogens.

Biosynthesis of glucosyl monophosphoryl undecaprenol and its role in lipoteichoic acid biosynthesis.

A glucophospholipid was detected in an incubation mixture containing UDP-glucose, MgCl2, ATP, and a particulate enzyme prepared from Streptococcus sanguis. The synthesis of this lipid was inhibited strongly by UDP and moderately by UMP. The molar ratio of glucose to phosphate in the purified lipid was found to be 1:1. Glucose and glucose 1-phosphate were released by mild alkaline hydrolysis of the glucophospholipid. The lipid produced by mild acid degradation of the purified lipid yielded a thin-layer chromatographic profile similar to that of acid-treated undecaprenol. One of the minor components exhibited the same mobility as untreated undecaprenol. To characterize further the lipid moiety of the glucophospholipid, a polyisoprenol was purified from the neutral lipid of S. sanguis. The polyisoprenol was converted in the presence of ATP, UDP-glucose, and the particulate enzyme into a lipid which exhibited the same thin-layer chromatographic mobility as the glucophospholipid. The structure of the polyisoprenol was determined by nuclear magnetic resonance and mass spectrometry to be an undecaprenol with an internal cis-trans ratio of 7:2. These results indicate that the glucophospholipid is glucosyl monophosphoryl undecaprenol. The glucosyl moiety of the glucophospholipid was shown to be incorporated in the presence of the particulate enzyme into a macromolecule which was characterized as a lipoteichoic acid by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and DEAE-cellulose column chromatography. This result indicates that glucosyl monophosphoryl undecaprenol is the direct glucosyl donor in the synthesis of lipoteichoic acid.

The genomic organization, complete mRNA sequence, cloning, and expression of a novel human intracellular membrane-associated calcium-independent phospholipase A(2).

During the sequencing of the long arm of chromosome 7 in the Human Genome Project, a predicted protein product of 40 kDa was identified, which contained two approximately 10-amino acid segments homologous to the ATP and lipase consensus sequences present in the founding members of a family of calcium-independent phospholipases A(2). Detailed inspection of the identified sequence (residues 79, 671-109,912 GenBank accession no. AC005058) demonstrated that it represented only a partial sequence of a larger undefined polypeptide product. Accordingly, we identified the complete genomic organization of this putative phospholipase A(2) through analyses of previously published expressed sequence tags, PCR of human heart cDNA, and 5'-rapid amplification of cDNA ends. Polymerase chain reaction and Northern blotting demonstrated a 3.4-kilobase message, which encoded a polypeptide with a maximum calculated molecular weight of 88476.9. This 3.4-kilobase message was present in multiple human parenchymal tissues including heart, skeletal muscle, placenta, brain, liver, and pancreas. Cloning and expression of the protein encoded by this message in Sf9 cells resulted in the production of two proteins of apparent molecular masses of 77 and 63 kDa as assessed by Western analyses utilizing immunoaffinity-purified antibody. Membranes from Sf9 cells expressing recombinant protein released fatty acid from sn-2-radiolabeled phosphatidylcholine and plasmenylcholine up to 10-fold more rapidly than controls. The initial rate of fatty acid release from the membrane fraction was 0. 3 nmol/mg.min. The recombinant protein was entirely calcium-independent, had a pH optimum of 8.0, was inhibited by (E)-6-(bromomethylene)-3-(1-naphthalenyl)-2H-tetrahydropyran-2-one (IC(50) = 3 microM), and was predominantly present in the membrane-associated fraction. Collectively, these results describe the genomic organization, complete mRNA sequence, and sn-2-lipase activity of a novel intracellular calcium-independent membrane-associated phospholipase A(2).

Long-term follow-up of generalized anxiety disorder.

The diagnostic stability and long-term prognosis of generalized anxiety disorder (GAD) remain the subjects of considerable controversy. We report the results of an investigation of the long-term outcome of an original sample of 44 subjects who participated in a medication trial. Subjects were reinterviewed approximately 16 months after completion of the study, using structured interviews. Fifty percent continued to fulfill criteria for GAD. Other concurrent axis I diagnoses were as follows: dysthymia, 11%; major depression, 7%; and social phobia, 7%. Regarding axis II comorbidity, subjects with chronic GAD were more likely to fulfill criteria for one or more personality disorders, especially in clusters B and C. In addition, follow-up subjects with GAD and with remitted GAD reported a statistically equivalent number of recent life events, although subjects with chronic GAD were more likely to report significant dissatisfaction with life. The findings indicate that although many subjects with GAD do not follow a chronic course, many others remain symptomatic. The results also suggest that GAD symptoms are not simply the result of a subject's recent negative experiences, and that life satisfaction measures are an accurate reflection of GAD outcome.

Hepatic extraction of organic anions in the rat depends on ligand hydrophobicity.

Non-bile-salt cholephilic organic anions are efficiently taken up by the liver. Recent work from our group has suggested the possible importance of relative hydrophobicity among various organic anions in hepatic uptake. To further validate and clarify this, we studied hepatic extraction of five different cholephilic dyes using the isolated perfused rat liver in single-pass mode. Albumin binding affinities and capacities for each of the ligands were measured in vitro to permit evaluation of in vivo interactions for each of them over a spectrum of unbound ligand concentrations. As expected, a strong positive correlation was found between ligand hydrophobicity and the relative degree of albumin binding affinity and capacity. Using appropriate experimental conditions, we also found a strong positive correlation between hepatic extraction efficiency for a given ligand and both its hydrophobicity and its unbound concentration. These data indicate that where the unbound ligand concentration is significant, the greater the ligand hydrophobicity, the greater is its efficiency of hepatic extraction. We conclude that hepatic extraction efficiency for non-bile-salt cholephilic organic anions depends on a combination of ligand hydrophilic/hydrophobic balance and the availability of the unbound ligand for uptake.

Identification of the calmodulin-binding domain of recombinant calcium-independent phospholipase A2beta. implications for structure and function.

Calcium-independent phospholipase A(2) (iPLA(2)) is the major phospholipase A(2) activity in many cell types, and at least one isoform of this enzyme class is physically and functionally coupled to calmodulin (CaM) in a reversible calcium-dependent fashion. To identify the domain in recombinant iPLA(2)beta (riPLA(2)beta) underlying this interaction, multiple techniques were employed. First, we identified calcium-activated CaM induced alterations in the kinetics of proteolytic fragment generation during limited trypsinolysis (i.e. CaM footprinting). Tryptic digests of riPLA(2)beta (83 kDa) in the presence of EGTA alone, Ca(+2) alone, or EGTA and CaM together resulted in the production of a major 68-kDa protein whose kinetic rate of formation was specifically attenuated in incubations containing CaM and Ca(+2) together. Western blotting utilizing antibodies directed against either the N- or C-terminal regions of riPLA(2)beta indicated the specific protection of riPLA(2)beta by calcium-activated CaM at a cleavage site approximately 15 kDa from the C terminus. Moreover, calcium-activated calmodulin increased the kinetic rate of tryptic cleavage near the active site of riPLA(2)beta. Second, functional characterization of products from these partial tryptic digests demonstrated that approximately 90% of the 68-kDa riPLA(2)beta tryptic product (i.e. lacking the 15-kDa C-terminus) did not bind to a CaM affinity matrix in the presence of Ca(2+), although >95% of the noncleaved riPLA(2)beta as well as a 40-kDa C-terminal peptide bound tightly under these conditions. Third, when purified riPLA(2)beta was subjected to exhaustive trypsinolysis followed by ternary complex CaM affinity chromatography, a unique tryptic peptide ((694)AWSEMVGIQYFR(705)) within the 15-kDa C-terminal fragment was identified by RP-HPLC, which bound to CaM-agarose in the presence but not the absence of calcium ion. Fourth, fluorescence energy transfer experiments demonstrated that this peptide (694) bound to dansyl-calmodulin in a calcium-dependent fashion. Collectively, these results identify multiple contact points in the 15-kDa C terminus as being the major but not necessarily the only binding site responsible for the calcium-dependent regulation of iPLA(2)beta by CaM.

Analytical gradient polyacrylamide gel-crossed line immunoelectrophores. A technique for locating species specific Mycobacterium tuberculosis H37RV antigens in polyacrylamide gel columns.

Analytical gradient polyacrylamide gel electrophoresis of antigens from Mycobacterium tuberculosis H37RV was combined with crossed line immunoelectrophoresis to locate the position of antigens in polyacrylamide gel columns that appeared to be specific for M. tuberculosis H37RV. Specificity was established by comparing H37RV with M. kansaii CE in the electrophoretic procedure and the relative mobility (Rm) values are being used in an effort to recover and enrich specific antigens from preparative gel columns. Such fractions should be more suitable in efforts to recover purified monospecific components from M. tuberculosis H37RV.