Fibroblast subpopulations in intra-oral wound healing

The objective of this study was to characterize fibroblasts at sequential time points during intra-oral wound healing in the rat. Experimental wounds were made at several time points in the mucoperiosteum of the palate of 35-day-old Wistar rats. Fibroblasts were cultured from the biopsies under standard conditions for the same number of passages. The expression of the integrin subunits alpha 1, alpha 6, and beta 1; and the intermediate filaments alpha-smooth muscle actin and vimentin were analyzed by flow cytometry. Western blot analysis was performed at 0, 8, and 60 days postwounding to confirm the expression of both intermediate filaments. The phenotypic profiles of fibroblasts cultured from subsequent stages in the wound healing process differed considerably. We conclude that distinct fibroblast phenotypes can be isolated from different stages in wound healing. These phenotypes remained stable during in vitro culturing. In addition, cryosections of the wound areas were made at identical time points and were immunohistochemically stained for the same antigens. The immunohistochemical staining correlated well to the flow-cytometric data. These results suggest the occurrence of multiple subpopulations of fibroblasts with a specialized function during wound healing. We hypothesize that undesirable consequences of wound healing might be prevented through the modulation of specific fibroblast subpopulations.     

Stress proteins as inducers and targets of regulatory T cells in arthritis

Immunization with microbial or mammalian stress proteins or heat-shock proteins in models of experimental autoimmunity has been observed to lead to increased disease resistance. Furthermore, such immunization has been proposed to result in the induction and expansion of T cells that suppress disease upon transfer. Comparisons of microbial heat-shock proteins with other conserved immunogenic proteins of bacterial origin have indicated a unique capacity for heat-shock proteins to induce a regulatory phenotype in T cells, such as reflected by the production of IL10. Also, studies in children with chronic arthritis have indicated that T-cell responses to heat-shock proteins are associated with a benign course of the disease and with remission. Furthermore, in patients, heat-shock-protein-(HSP-) activated T cells were shown to display regulatory phenotypes consistent with CD4+ CD25+ T regulatory cells.     

Multicenter quality assessment of PCR methods for detection of enteroviruses

We conducted a multicenter evaluation of commercial and in-house PCR methods for the detection of enteroviruses. Three coded panels of test and control RNA samples, artificial clinical specimens, and representative enterovirus serotypes were used to assess amplification methods, RNA extraction methods, and reactivities with different enterovirus serotypes. Despite several differences between PCR methods, there was good agreement, although some variation in sensitivity was observed. Most PCR methods were able to detect enterovirus RNA derived from 0.01 50% tissue culture infective dose (TCID50) and were able to detect at least 1 TCID50 of enterovirus in cerebrospinal fluid, stool, or throat swab specimens. Most were also able to detect a wide range of enterovirus serotypes, although serotypic identification was not possible. Some laboratories experienced false-positive results due to PCR contamination, which appeared to result mainly from cross-contamination of specimens during RNA extraction. Provided that this problem is overcome, these PCR methods will prove to be a sensitive and rapid alternative to cell culture for the diagnosis of enterovirus infection.     

High-density lipoprotein cholesterol subfractions in chronic uremia

Cholesterol content in high-density lipoprotein (HDL) subfractions has been studied in 108 patients at different evolutive stages of chronic renal failure (CRF) under conservative treatment. Results have been compared with healthy control subjects, patients receiving hemodialysis, and renal graft recipients. Significant low levels of total HDL and HDL2 cholesterol are observed in men with CRF. The more severe the CRF, the more likely that total HDL and HDL2 cholesterol will be low. Moreover, a significant inverse correlation is found between HDL2 cholesterol and serum creatinine levels. In women, although a decrease in total and HDL2 subfraction is observed, no significant differences are found across the severity of CRF. Serum HDL2 cholesterol levels are decreased in men and women receiving hemodialysis, while raised total HDL and HDL2 cholesterol levels are observed in normally functioning renal grafts. These results indicate that according to the "HDL hypothesis," despite other associated risk factors, the high cardiovascular mortality rates noted mainly in men with CRF under conservative treatment and in patients receiving hemodialysis could be explained, at least in part, by the sustained and progressive decrease in total HDL and HDL2 values. From this point of view, our study suggests the need to promote early kidney transplantation.     

Drug Delivery in Pediatric Patients with Asthma

This review discusses published evidence on the use of metered dose inhaler (MDI)/spacer combinations and nebulizers for delivering β₂ adrenoeeptor agonists (β₂ agonists) in stable and acute severe childhood asthma. Although nebulizers have been the mainstay of inhalation therapy in childhood asthma for many years, these devices are cumbersome, bulky, time-consuming, and expensive to use. As a result, over the past decade the emphasis of inhalation therapy in children has shifted from nebulizers to metered-dose inhalers (MDI) in combination with spacer devices. MDI/spacer combinations have been shown to be a practical and effective way of delivering medications for inhalation in children with asthma, irrespective of their age. Lung deposition increases with age, being low in young children. This underscores the need for administering relatively high nominal dosages of medication when using an MDI/spacer in young children; the most practical approach is to use the same dose, irrespective of the child’s age or weight. In children with both stable chronic asthma and acute severe asthma, numerous studies have shown that β₂ agonists can be delivered by MDI/spacer at least as effectively and safely as by nebulizer. Although nebulizers may still play a role in status asthmaticus (because they allow combined administration of β₂ agonists, anticho-linergic agents, and oxygen in a single procedure), the vast majority of children of all ages with acute severe asthma can be managed effectively and safely by β₂ agonists delivered via MDI/spacer. This cost-effective and practical approach should, therefore, be preferred.     

Peptide-induced nasal tolerance for a mycobacterial heat shock protein 60 T cell epitope in rats suppresses both adjuvant arthritis and nonmicrobially induced experimental arthritis

Adjuvant arthritis (AA) can be induced in Lewis rats by immunization with mycobacterial antigens. Passive transfer of a T cell clone recognizing the 180–188 amino acid sequence in mycobacterial heat shock protein 60 (hsp60) was found to induce AA. In the present study, we investigated whether tolerance was obtained for this AA-associated T cell epitope after intranasal or s.c. administration of a peptide containing this epitope. Two 15-mer peptides containing the mycobacterial hsp60 sequences 176–190 and 211–225 were used; 176–190 contained the T cell epitope 180–188, which was recognized by the arthritogenic T cell clone A2b and was the immunodominant hsp60 T cell epitope after induction of AA, and 211–225 contained a T cell epitope that was recognized both after induction of arthritis with whole Mycobacterium tuberculosis and after immunization with mycobacterial hsp60. In rats treated intranasally or subcutaneously with 176–190 and immunized with mycobacterial hsp60, proliferative responses to 176–190 were reduced. Proliferative responses to 211–225 and to whole mycobacterial hsp60 were not affected. AA was inhibited intranasally in the 176–190-treated rats but not in the 211–225-treated rats. Moreover, intranasal 176–190 led to similar arthritis-protective effects in a nonmicrobially induced experimental arthritis (avridine-induced arthritis). Therefore, tolerance for a disease-triggering, microbial cartilage-mimicking epitope may cause resistance to arthritis irrespective of the actual trigger leading to development of the disease.