Sarcomeric determinants of striated muscle relaxation kinetics

Ca²⁺ is the primary regulator of force generation by cross-bridges in striated muscle activation and relaxation. Relaxation is as necessary as contraction and, while the kinetics of Ca²⁺-induced force development have been investigated extensively, those of force relaxation have been both studied and understood less well. Knowledge of the molecular mechanisms underlying relaxation kinetics is of special importance for understanding diastolic function and dysfunction of the heart. A number of experimental models, from whole muscle organs and intact muscle fibres down to single myofibrils, have been used to explore the cascade of kinetic events leading to mechanical relaxation. By using isolated myofibrils and fast solution switching techniques we can distinguish the sarcomeric mechanisms of relaxation from those of myoplasmic Ca²⁺ removal. There is strong evidence that cross-bridge mechanics and kinetics are major determinants of the time course of striated muscle relaxation whilst thin filament inactivation kinetics and cooperative activation of thin filament by cycling, force-generating cross-bridges do not significantly limit the relaxation rate. Results in myofibrils can be explained well by a simple two-state model of the cross-bridge cycle in which the apparent rate of the force generating transition is modulated by fast, Ca²⁺-dependent equilibration between off- and on-states of actin. Inter-sarcomere dynamics during the final rapid phase of full force relaxation are responsible for deviations from this simple model.     

Acute graft-versus-host disease and steroid treatment impair CD11c+ and CD123+ dendritic cell reconstitution after allogeneic peripheral blood stem cell transplantation.

Human dendritic cells (DC) comprise 2 subsets-plasmacytoid CD123(+) and myeloid CD11c(+) DC-that may have distinct roles in the regulation of immunity after allogeneic hematopoietic stem cell transplantation. In this study, we analyzed the kinetics of CD123(+) DC and CD11c(+) DC reconstitution in 31 patients who underwent transplantation with allogeneic granulocyte colony-stimulating factor-mobilized peripheral blood (PB) stem cells from HLA-identical sibling donors after myeloablative conditioning. Lineage marker-negative HLA-DR(+) CD11c(+) CD11c(+) DC and lineage marker-negative HLA-DR(+) CD123(+) CD123(+) DC, as well as monocytes and lymphoid subsets, were enumerated in donor grafts and in the PB of patients at various time points after transplantation. Reconstitution of both CD11c(+) DC and CD123(+) DC to normal levels occurred within 6 to 12 months and was not affected by the diagnosis, preparatory regimen, or graft composition. However, PB CD11c(+) DC and CD123(+) DC counts were significantly reduced in patients with acute GVHD grade II to IV (at 1 and 3 months) and grade I (at 1 month). Patients with chronic GVHD instead showed reduced CD123(+) DC counts only 6 months after transplantation. Moreover, treatment with steroids (>0.1 mg/kg) was significantly associated with reduced PB CD11c(+) DC and CD123(+) DC counts at all time points after transplantation. In multivariate analysis, only acute GVHD affected DC reconstitution early after transplantation. These results will prompt new studies addressing whether DC reconstitution correlates with immunity against infectious agents or with graft-versus-tumor reactions after PB stem cell allotransplantation.     

From surnames to the history of Y chromosomes: the Sardinian population as a paradigm

A total of 202 Sardinian male subjects were examined for 13 biallelic stable markers, the complex 49a,f/TaqI system and three microsatellites of the Y chromosome in order to investigate, through surname analysis, on a possible territorial heterogeneity inside the island. The study of geographical distribution and linguistic derivation of Sardinian surnames allow us to discover their 'probable place of origin' and reconstruct ancient genetic isolates which borders are, today, no more recognizable. The molecular analysis revealed that about 90% of the Sardinian Y chromosomes fell into haplogroups E-M35, G-M201, I-M26, J-12f2 and R-M269. In contrast with the territorial homogeneity of these haplogroups, when the individuals were distributed according to their birthplace, a significant difference between the three historically and culturally distinct geographical areas into which Sardinia can be subdivided was observed when the individuals were distributed according to the ancestral location of surnames. In particular, the major contribution to this heterogeneity is due to the 'Sardinian-specific' haplogroup I-M26 (almost completely associated with the 49a,f-Ht12/12f2-10Kb/YCAIIa-21/YCAIIb-11 compound haplotype), which shows both a significantly higher incidence in the central-eastern (archaic) area and a significantly lower frequency in the northern area. The results of this study agree with the hypothesis that the ancestral homeland of this specific subset of haplogroup I is the mountainous central-eastern area of Sardinia, where the population underwent a long history of isolation since ancient times, and highlight the informative power of the surname analysis.     

Isolation of human immunodeficiency virus (HIV) from plasma during primary HIV infection

Human immunodeficiency virus (HIV) has been isolated from plasma in 6 of 7 patients showing clinical symptoms of a primary HIV infection. Parallel cultures from peripheral blood mononuclear cells (PBMC) yielded virus in 5 patients. In one case, virus could only be isolated from the cerebrospinal fluid but not from peripheral blood. Detectable viremia was transient and preceded the appearance of HIV specific antibodies. After cessation of acute symptoms, the frequency of HIV isolations was similar to that of asymptomatic carriers (23 and 26%, respectively). The role of the immune response in terminating detectable viremia remains to be established.     

Phagocytosis in the colonial ascidian Botryllus schlosseri

Phagocytosis by Botryllus schlosseri hemocytes in influenced by temperature, pH, concentration, and physicochemical properties of the test particles and requires Ca²⁺ or Mg²⁺ ions to occur. Phagocytes recognized glucosyl or mannosyl residues on the surface of yeast cells, and a respiratory burst is associated with phagocytosis, as indicated by increased superoxide production. Factors that enhance phagocytosis of yeast, sheep red blood cells, and latex beads and reduce the uptake of yeast and sheep erythrocytes are present in the plasma.     

Biodegradation of poly(D(–)-3-hydroxybutyrate)/atactic poly(epichlorohydrin) blends by aureobacterium saperdae

The biodegradability of solution-cast films of poly(D (–)-3-hydroxybutyrate) (PHB) blended with the melt-compatible component atactic poly(epichlorohydrin) (aPECH) was investigated. A bacterium which produced extracellular enzymes that degraded PHB even when blended with aPECH was isolated, and tentatively designated as Aureobacterium saperdae. The growth rate of A. saperdae decreased with increasing aPECH content in the blend, up to films containing 60 wt.-% aPECH, at which composition growth was completely inhibited. The decrease in the bacterial growth rate could be due to the dilution of PHB molecules on the blend film surface caused by the presence of aPECH molecules. At the stationary phase of bacterial growth the percentage of weight loss of blend films decreased with increasing aPECH fraction, which was probably due to the lower accessibility of PHB when blended with aPECH. During the bacterial growth only PHB was metabolized, whereas neither degradation nor abiotic release of aPECH was detected for blend films.     

Absence of coding mutations in the X-linked genes neuroligin 3 and neuroligin 4 in individuals with autism from the IMGSAC collection.

Neuroligin abnormalities have been recently implicated in the aetiology of autism spectrum disorders (ASD), given the finding of point mutations in the two X-linked genes NLGN3 and NLGN4X and the important role of neuroligins in synaptogenesis. To enquire on the relevance and frequency of neuroligin mutations in ASD, we performed a mutation screening of NLGN3 and NLGN4X in a sample of 124 autism probands from the International Molecular Genetic Study of Autism Consortium (IMGSAC). We identified a new non-synonymous variant in NLGN3 (Thr632Ala), which is likely to be a rare polymorphism. Our data indicate that coding mutations in these genes are very rarely associated to ASD.     

FAS and FAS-L expression by tumor cells and lymphocytes in breast carcinomas and their lymph node metastases

FAS receptor (FAS, CD95) and FAS ligand (FAS-L, CD95-L) are complementary members of a particular apoptotic pathway that plays a major role in immune regulation. The activation of FAS-L may trigger cytotoxic mechanisms leading to the death of FAS-expressing cells. Tumor cells and tumor-infiltrating lymphocytes (TIL) may express FAS and FAS-L in various proportions, and their interplay may affect tumor behavior. In the present study, we explored the expression of FAS and FAS-L in 28 mammary carcinomas (19 ductal and 9 lobular) and in their lymph node metastases. The expression of these mediators in immunostained sections was graded and evaluated comparatively between normal and neoplastic mammary epithelium, between tumor cells and TILs, and between mammary carcinoma cells and their lymph node metastases. We demonstrated the coexpression of FAS and FAS-L by breast carcinoma cells and TIL, with FAS expressed more strongly by normal epithelial cells and TIL than tumor cells. FAS-L was better stained on tumor cells than on TIL. There was equal or greater expression of FAS and FAS-L in the primary tumors and their TIL than in the metastatic counterparts. Comparing the expression of FAS with that of FAS-L, we recorded FAS equal or stronger than FAS-L in the primary mammary tumors and the reversal of their expression, FAS-L greater than FAS in the lymph node metastases. These results are consistent with reports of studies with other tumors, suggesting that the upregulated FAS-L indicates an increased ability of tumor cells to induce apoptosis in TIL and in the normal tissues invaded. However, it is understood that the FAS/FAS-L system, although essential for apoptosis, is only a contributing factor to the complex process of tumor invasion and antitumor defense.     

The nature of high frequency sister chromatid exchange cells (HFCs)

We employed the three-way differential staining technique (TWD),which allows SCEs to be distinguished on a per generation basisby scoring third metaphases (M3), in order to study the spontaneouslevels of SCEs in normal and high frequency cells (HFCs) thatoccurred in the first (S1), second (S2) and third (S3) S phases.Fifty one of 900 lymphocytes from 37 healthy donors were definedas HFCs by calculating the 95th percentile of the distributionof SCEs in S1 + S2. ‘Normal’ cells presented almostthe same number of SCEs after the first, second and third cellcycles (SCE averages of 2.43, 2.04 and 3.53 respectively). Incontrast, HFCs showed a higher SCE count in SI, which decreasedrapidly through the cycles and reached baseline level at S3(SCE averages of 7.18, 4.29 and 3.45 respectively). This wouldsuggest that the lesions responsible for the higher SCE frequencyin HFCs were effectively removed after two cell cycles and stronglysupport the hypothesis that HFCs are lymphocytes which accumulatehigher levels of DNA lesions through time. ¹To whom correspondence should be addressed     

Could mitochondrial haplogroups play a role in sporadic amyotrophic lateral sclerosis?

Mitochondrial impairment has been implicated in the pathogenesis of the amyotrophic lateral sclerosis (ALS). Furthermore, mitochondrial-specific polymorphisms were previously related to other neurodegenerative diseases, such as Parkinson, Friedreich and Alzheimer disease. To investigate if specific genetic polymorphisms within the mitochondrial genome (mtDNA) could act as susceptibility factors and contribute to the clinical expression of sporadic ALS (sALS), we have genotyped predefined European mtDNA haplogroups in 222 Italian patients with sALS and 151 matched controls. Individuals classified as haplogroup I demonstrated a significant decrease in risk of ALS versus individuals carrying the most common haplogroup, H (odds ratio 0.08, 95% confidence interval 0.04-0.4, p < 0.01). Further stratification of the dataset by sex, age and site of onset of disease and survival failed to reach significance for association. Our study provides evidence of the contribution of mitochondrial variation to the risk of ALS development in Caucasians. Further it may help elucidate the mechanism of the mitochondrial dysfunction detectable in ALS, and may be of relevance in development of strategies for the treatment of this disease.