A T-cell epitope encoded by a subset of HLA-DPB1 alleles determines nonpermissive mismatches for hematologic stem cell transplantation

The importance of HLA-DPB1 matching for the outcome of allogeneic hematologic stem cell (HSC) transplantation is controversial. We have previously identified HLA-DPB1*0901 as a target of cytotoxic T cells mediating in vivo rejection of an HSC allograft. Here we show that HLA-DPB1*0901 encodes a T-cell epitope shared by a subset of DPB1 alleles that determines nonpermissive mismatches for HSC transplantation. Several T-cell clones obtained from the patient at the time of rejection showed HLA-DP restricted recognition of allogeneic targets expressing HLA-DPB1*0901, *1001, *1701, *0301, *1401, and *4501, but not other alleles. Based on these findings, we developed an algorithm for prediction of nonpermissive HLA-DPB1 mismatches. Retrospective evaluation of 118 transplantations showed that the presence of nonpermissive HLA-DPB1 mismatches was correlated with significantly increased hazards of acute grade II to IV graft-versus-host disease (HR = 1.87, P =.046) and transplantation-related mortality (HR = 2.69, P =.027) but not relapse (HR = 0.98, P =.939), as compared with the permissive group. There was also a marked but statistically not significant increase in the hazards of overall mortality (HR = 1.64, P =.1). These data suggest that biologic characterization of in vivo alloreactivity can be a tool for definition of clinically relevant nonpermissive HLA mismatches for unrelated HSC transplantation.     

Chronic idiopathic myelofibrosis: independent prognostic importance of bone marrow microvascular density evaluated by CD105 (endoglin) immunostaining.

Microvascular density (MVD) is substantially increased in bone marrow biopsies of patients with chronic idiopathic myelofibrosis (CIMF). CD105, a useful molecule for assessing MVD in various malignancies, is preferentially expressed by recently formed microvessels. Increased serum-soluble CD105 in patients with chronic myeloproliferative disorders, including CIMF, was documented. CD105 MVD has not so far been investigated in CIMF: to this end, the results in 55 patients with CIMF and 21 controls were compared with the conventional CD34 immunostaining as well as traditional histological and clinical disease features. The MVD mean values estimated by both CD105 and CD34 were significantly higher in CIMF patients than in controls (P<0.00001). In addition, the proportion of CD105-positive megakaryocytes was significantly higher in CIMF than in controls (P<0.0001). A degree of reticulin fibrosis >2 correlated with increased CD105 MVD (P=0.05). A multivariate analysis confirmed that CD105-positive MVD was an independent adverse prognosticator. This study demonstrates that while MVD, as assessed by both CD34 and CD105 immunostaining, is significantly increased in CIMF, only CD105-determined MVD correlates with the degree of fibrosis and is prognostically relevant. These findings provide a rationale for the investigational use of anti-CD105-targeted drugs in CIMF.     

Abnormal DNA content in liver-cell dysplasia: a flow cytometric study

To clarify the true nature of liver-cell dysplasia (LCD), a flow cytometric study has been performed. The DNA content of hepatocytes from 26 cases of cirrhosis with diffuse areas of LCD was investigated and compared to that of hepatocytes from 21 control patients with non-neoplastic and neoplastic liver conditions. Flow cytometric analysis was performed on propidium-stained nuclei from archival paraffin-embedded material. Analysis was directed to assessment of diploid as well as non-diploid peaks by calculation of DNA index (DI), using normal hepatocytes present in each sample as individual and specific references. Since only samples containing at least 10,000 nuclei were considered suitable for analysis, 4 of the 26 LCD cases were discarded. Eight of 22 LCD cases had an abnormal DNA content compared with 0/11 non-neoplastic cases (p less than 0.05) and 8/10 hepatocellular carcinomas (p less than 0.05). Non-neoplastic control cases displayed uniformly diploid stemlines whereas hepatocellular carcinomas had in 8/10 cases bimodal or trimodal populations. Thus, LCD is a heterogeneous lesion in terms of ploidy, and the abnormal DNA content observed in some cases supports its pre-neoplastic nature.     

Vaccination recommendations for patients with neuromuscular disease

Neuromuscular diseases (NMDs) encompass a broad spectrum of conditions. Because infections may be relevant to the final prognosis of most NMDs, vaccination appears to be the simplest and most effective solution for protecting NMD patients from vaccine-preventable infections. However, very few studies have evaluated the immunogenicity, safety, tolerability, and efficacy of different vaccines in NMD patients; therefore, detailed vaccination recommendations for NMD patients are not available. Here, we present vaccination recommendations from a group of Italian Scientific Societies for optimal disease prevention in NMD patients that maintain high safety levels. We found that NMD patients can be classified into two groups according to immune function: patients with normal immunity and patients who are immunocompromised, including those who intermittently or continuously take immunosuppressive therapy. Patients with normal immunity and do not take immunosuppressive therapy can be vaccinated as healthy subjects. In contrast, immunocompromised patients, including those who take immunosuppressive therapy, should receive all inactivated vaccines as well as influenza and pneumococcal vaccines; these patients should not be administered live attenuated vaccines. In all cases, the efficacy and long-term persistence of immunity from vaccination in NMD patients can be lower than in normal subjects. Household contacts of immunocompromised NMD patients should also be vaccinated appropriately.     

Granulomatous and nongranulomatous lymphadenitis in sarcoidosis. An immunophenotypic study of seven cases

Five cases of prescalenic granulomatous lymphadenitis (GL) and 2 cases of prescalenic nongranulomatous lymphadenitis (NGL) in 7 patients with sarcoidosis were studied with a large panel of monoclonal antibodies (anti-CD 1, CD 3, CD 4, CD 8, CD 14, CD 22, CD 25, HLA-DR, HLA-DQ, HNK-1, R 4/23). Immunopathologic analysis was performed by studying three different compartments of GL--granuloma, intergranulomatous area and sinuses--and of NGL--cortex, paracortex and sinuses. Intra- and intergranulomatous T lymphocytes were mainly of the CD 4 type in 4 out of 5 cases; in all the cases less than 25% of T lymphocytes stained also for CD 25. Epithelioid cells were HLA-DR+, HLA-DQ+, CD 14+ and, frequently, CD 1+ and CD 25+, the latter positivity being mainly restricted to the marginally located epithelioid cells. Sinuses were filled with T- and B- cells; sinusal histiocytes were HLA-DR+, HLA-DQ+, CD 14+ and, frequently, CD 1+ and CD 25+. In the cortex and paracortex of NGL, T-cell subsets paralleled the distribution and ratio observed in the intergranulomatous area of GL; furthermore the immunophenotype of NGL sinusal histiocytes roughly overlapped that observed in the same district of GL with a strong CD 1 and CD 25 positivity. These results, besides confirming the global imbalance of the CD 4/8 ratio in all the areas of GL, seem to demonstrate that the prevalence of CD 4+ or CD 8+ T-cells probably reflects different functional phases of the granulomatous process.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of recombinant human stem cell factor (rh-SCF) on colony formation and long-term bone marrow cultures (LTBMC) in patients with myelodysplastic syndromes

Abstract: Stem cell factor (SCF), the ligand of the c-kit receptor, is a potent enhancing cytokine for haematopoietic cells in the presence of IL-3, GM-CSF and erythropoietin (Epo). In the clonogenic assays of 63 MDS patients, the addition of rh-SCF + GM-CSF and/or IL-3 induced a significant increase (p<0.001) in the number and size of CFU-GM. Never reaching the levels of controls, this increase was seen in all FAB subtypes, but particularly in RA. There was no significant increase in cluster formation, even in RAEB or RAEBt. Rh-SCF (10 ng/ml) led to mean increases of up to 26 times in the number of Epo-dependent BFU-E colonies, particularly in RA (p<0.001) and RAEB (p<0.05). Individual responses varied widely (especially in RA) from no response to supranormal levels. Added to the weekly refeed of 37 MDS LTBMC, SCF (10 ng/ml) induced only a 7% mean increase in both cell output and the number of clonogenic cells recovered in the supernatant. Immunohistochemical examination of the supernatant showed significant increases in differentiating myeloid cells in all examined cases, and in erythroid cells in 3 cases; blast cells increased in only 3 cases. These data suggest that rh-SCF is capable of at least partially reversing defective MDS myeloid haematopoiesis, and leads no overt risk of leukaemic transformation. Its potent effect on erythroid cells is encouraging for future clinical applications in patients, particulary if they are selected by means of in vitro tests.     

The novel mitochondrial tRNAAsn gene mutation m.5709T>C produces ophthalmoparesis and respiratory impairment

Although mutations in mitochondrial tRNAs constitute the most common mtDNA defect, the presence of pathological variants in mitochondrial tRNA(Asn) is extremely rare. We were able to identify a novel mtDNA tRNA(Asn) gene pathogenic mutation associated with a myopathic phenotype and a previously unreported respiratory impairment. Our proband is an adult woman with ophthalmoparesis and respiratory impairment. Her muscle biopsy presented several cytochrome c oxidase-negative (COX-) fibres and signs of mitochondrial proliferation (ragged red fibres). Sequence analysis of the muscle-derived mtDNA revealed an m.5709T>C substitution, affecting mitochondrial tRNA(Asn) gene. Restriction-fragment length polymorphism analysis of the mutation in isolated muscle fibres showed that a threshold of at least 91.9% mutated mtDNA results in the COX deficiency phenotype. The new phenotype further increases the clinical spectrum of mitochondrial diseases caused by mutations in the tRNA(Asn) gene.