Lígia M. I. Fukumori

Imunofluorescência direta e indireta

Immunofluorescence is a valuable auxiliary diagnostic tool for autoimmune bullous diseases and inflammatory disorders, since their clinical and histopathologic findings may be inconclusive. It is a feasible laboratory method that requires experienced technicians and detects in situ and circulating immune deposits that may be involved in the pathogenesis of such skin diseases.

The elevated interferon gamma production is an important immunological marker in HAM/TSP pathogenesis.

Human T‐lymphotropic virus type 1 (HTLV‐1) is the agent of the HTLV‐1‐associated myelopathy/tropical spastic paraparesis (HAM/TSP), which may occur in >5% of patients during their lifetime. HTLV‐1‐infection causes disturbances in the immune system, and the viral load may also play an important role in the pathogenesis of HAM/TSP. Some cytokines are involved in the pathogenesis of this disorder. We have determined IL‐2, IL‐4, IL‐10, IL‐12 p70, IFN‐γ and TNF‐α production among HTLV‐1‐infected subjects from our HTLV‐out Clinic in Institute of Infectious ‘Emílio Ribas’ in Sao Paulo city, Brazil. PBMC obtained from healthy controls (n = 32), asymptomatic HTLV‐1 carriers (n = 68) and HAM/TSP patients (n = 44) were grown in the absence and in the presence of phytohaemagglutinin (PHA), and the supernatants’ fluids were measured for cytokines production. IL‐2 levels were increased in the asymptomatic HTLV‐1 carriers, and IFN‐γ was increased in both groups of patients (asymptomatic HTLV‐1 carriers and more significantly among HAM/TSP patients). IL‐4, IL‐10, TNF‐α and IL‐12 p70 levels were not significantly increased on both groups of patients, as compared with controls. The major finding of this study is that IFN‐γ was an important cytokine for the HAM/TSP pathogenesis. Therefore, immune modulation of IFN‐γ may be critical to treat of HAM/TSP patients.

Endemic pemphigus foliaceus (fogo selvagem) and pemphigus vulgaris: immunoglobulin G heterogeneity detected by indirect immunofluorescence

UNLABELLED Pemphigus are autoimmune intraepidermal blistering diseases in which immunoglobulin G (IgG) autoantibodies are directed against desmosomal glycoproteins. The aim of this study was to determine the IgG subclass profile of endemic pemphigus foliaceus (fogo selvagem) and pemphigus vulgaris utilizing indirect immunofluorescence. PATIENTS AND METHODS Twenty-five patients with pemphigus vulgaris, 25 with endemic pemphigus foliaceus (fogo selvagem), and 25 healthy controls were analyzed by indirect immunofluorescence for circulating autoantibodies (total IgG and its subclasses). RESULTS Our data revealed a significant correlation (P <.05) of disease activity and autoantibody levels in both forms of pemphigus, i.e., negative titers related to clinical remission, whereas positive results related to active disease. Immunoglobulin G subclass analysis in fogo selvagem demonstrated that in patients in remission, 56% showed positive immunoglobulin G4; in active disease, immunoglobulin G4 was the predominant subclass (100% positive in all cases). The IgG subclass profile in pemphigus vulgaris showed that in patients in remission, only 10% were positive for immunoglobulin G4; in active disease, positivity for immunoglobulin G4 was present in 78% to 88% of the cases. CONCLUSION Subclass characterization of immunoglobulin G autoantibodies is a useful tool for pemphigus follow-up, since immunoglobulin G4 (IgG4) is the subclass that is closely related to recognition of pathogenic epitopes, and consequently with disease activity. Careful monitoring should be performed for fogo selvagem in clinical remission with a homogeneous IgG4 response, since this may indicate more frequent relapses.

Human T‐cell lymphotropic virus type 1 infective dermatitis emerging in adulthood

Background  Infective dermatitis (ID) is a rare dermatologic condition of childhood that has been linked to human T‐cell lymphotropic virus type 1 (HTLV‐1).

Direct immunofluorescence in lupus erythematosus (LE)

One hundred and twenty-six patients with LE were studied. They were distributed as follows: 84 with DLE, 13 with SALE and 29 with SLE. Biopsies from the skin lesions were performed and submitted to DIF. Positive results were equal to 69, 61.5 and 72.4 percent of the DLE, SALE and SLE cases, respectively. These data are in accordance with the literature. IgM was the most frequently found immunoglobulin, followed by the association IgM + C3.

Imunomapeamento nas epidermólises bolhosas hereditárias

Immunological mapping, an immunofluorescence technique, is currently the method most used to diagnose and differentiate the principal types of hereditary epidermolysis bullosa, since this technique is capable of determining the level of cleavage of this mechanobullous disease.

Low DNA HTLV-2 proviral load among women in Sao Paulo City

BACKGROUND HTLV-2 infections are almost always asymptomatic, and diseases associated with the infection are rarely reported. Little information is available on the relationship between HTLV-2 proviral load and gender or expression of disease, especially among patients with HIV-1 co-infection. METHODS We studied 77 HTLV-2-infected subjects followed in our clinic for the last 9 years; 53 (69%) of them were co-infected with HIV-1. HTLV-2 DNA proviral load (PVL) was measured by real time PCR, a test with a sensitivity of 10 in 10(4) PBMCs. RESULTS Six of 53HTLV-2/HIV-1 cases had a myelopathy (all of them had undetectable PVL of HTLV-2). Only 3 of 35 women (2 out of 3 co-infected with HIV) had a detectable PVL, whereas 10 of 42 men had a detectable PVL. Regardless of their HIV status women had significantly lower PVL than men (10 vs. 43 copies/10(4) PBMCs, p<0.05). CONCLUSIONS We noticed the occurrence of myelopathy in HTLV-2/HIV-1 co-infected patients, with undetectable HTLV-2 viral load. There was a sex difference in viral load for HTLV-2, what may be the result in mode of transmission or acquisition of the virus.

Neither molecular diversity of the envelope, immunosuppression status, nor proviral load causes indeterminate HTLV western blot profiles in samples from human T-cell lymphotropic virus type 2 (HTLV-2)-infected individuals.

Although human T‐cell lymphotropic virus type 2 (HTLV‐2) is considered of low pathogenicity, serological diagnosis is important for counseling and monitoring. The confirmatory tests most used are Western blot (WB) and PCR. However, in high‐risk populations, about 50% of the indeterminate WB were HTLV‐2 positives by PCR. The insensitivity of the WB might be due to the use of recombinant proteins of strains that do not circulate in our country. Another possibility may be a high level of immunosuppression, which could lead to low production of virus, resulting in low stimulation of antibody. We found one mutation, proline to serine in the envelope region in the position 184, presented at least 1/3 of the samples, independent the indeterminate WB profile. In conclusion, we found no correlation of immune state, HTLV‐2 proviral load, or env diversity in the K55 region and WB indeterminate results. We believe that the only WB kit available in the market is probably more accurate to detect HTLV‐1 antibodies, and some improvement for HTLV‐2 detection should be done in the future, especially among high‐risk population. J. Med. Virol. 82: 837–842, 2010.

Patterns of In vitro Lymphoproliferative Responses Among HTLV-1-infected Subjects: Upregulation by HTLV-1 During HIV-1 Co-infection

The present study evaluated the in vitro response to different mitogens and a candidin antigen (CMA) in Human T‐cell lymphotropic virus type 1 (HTLV‐1) and co‐infected HIV‐1/HTLV‐1 patients, to identify if this co‐infection may modify the spontaneous lymph proliferative response. Peripheral blood mononuclear cells from 72 healthy seronegative controls, 75 asymptomatic HTLV‐1‐infected carriers, 42 HAM/TSP cases, 33 solely HIV‐1‐infected subjects and 24 HIV‐1/HTLV‐1 patients were assayed in the presence and absence of mitogens (PHA, PWM and OKT3) and CMA. The HAM/TSP group had the highest proliferation rate at 3 and 6 days after culture. HAM/TSP cases showed decreased response to PHA, compared with asymptomatic HTLV‐1 subjects, and most important, the co‐infected HIV‐1/HTLV‐1 cases presented a similar response to HTLV‐1‐infected subjects after 3 days of culture. The singles HIV‐1‐infected group had decreased in vitro response. It appears that during co‐infection, the HTLV‐1 regulatory proteins overwhelm the action of HIV‐1 regulatory proteins.