Recombinant 60-kDa heat shock protein from Paracoccidioides brasiliensis: is it a good antigen for serological diagnosis of paracoccidioidomycosis?

Paracoccidioides brasiliensis and P. lutzii are fungi that cause paracoccidioidomycosis (PCM), the most prevalent systemic mycosis in South America. For serological diagnosis, although 43-kDa glycoprotein (gp43) is regarded as highly specific for PCM, the occurrence of false negative reactions in sera from patients infected with P. lutzii suggests that preparation with only one antigen is not recommended. Heat shock proteins are feasible alternatives as a second antigen because they are often highly immunogenic. In this study, we evaluated the usefulness of recombinant 60-kDa heat shock protein from P. brasiliensis (rPbHsp60) for the serological diagnosis of PCM. Using western blotting assay, we observed that 77.3% of the sera from PCM patients were positive to rPbHsp60, with 90.9% positivity to recombinant gp43 (rgp43). More importantly, sera from healthy subjects had 27% positivity to rPbHsp60 and none to rgp43. When rPbHsp60 was used in ELISA, we did not observe significant differences between the reactions with sera from PCM patients and healthy subjects, while the difference was clearly evident when the antigen was rgp43. Furthermore, rPbHsp60 was recognized by sera from patients with histoplasmosis, aspergillosis, sporotrichosis or tuberculosis in an ELISA test. These results show that rPbHsp60 is not a good antigen for PCM diagnosis.

Evaluation of circulatory and salivary levels of heat shock protein 60 in periodontal health and disease.

Background: Self‑antigens such as heat shock protein 60 (HSP 60) have recently been implicated in the periodontal disease pathogenesis. There is scant evidence regarding HSP 60 levels in circulation and saliva following periodontal disease and its possible relation to systemic inflammation. Aim of the Study: The aim was to evaluate the circulatory and salivary levels of HSP 60 in periodontal health and disease and to correlate it with high sensitivity C‑reactive protein (hs‑CRP). Materials and Methods: Forty‑five peripheral blood samples were collected from two groups of patients (periodontally healthy ‑ Group A [22 patients] and periodontal disease ‑ Group B [23 patients]). Serum, cell lysates, and saliva samples were used to detect HSP 60 levels in both groups by enzyme linked immunosorbent assay technique. Measurement of hs‑CRP was performed using an immunoturbidimetric assay. Statistical analysis was done using the student t‑test and Pearson’s correlation. Results: Circulatory HSP 60 was significantly increased in periodontal disease compared to health (P ‑ 0.038). There was a significant correlation between the totals circulating HSP 60 and hs‑CRP (P ‑ 0.052), but there was no significant correlation between the salivary HSP 60 and hs‑CRP levels in periodontal disease. Conclusion: Circulating HSP 60 levels may play a role in the systemic inflammatory state produced by periodontal disease. Salivary HSP 60 may not be used as a surrogate to determine systemic inflammation.

High sensitivity C-reactive protein and immunoglobulin G against chlamydia pneumoniae and chlamydial heart shock protein-60 in ischemic heart disease

Inflammation and infectious agents such as Chlamydia pneumoniae have been associated with cardiovascular disease. To evaluate the serum high sensitivity C - reactive protein [hs-CRP] and antibodies against Chlamydia pneumoniae and Chlamydial heat shock protein-60 [Cp-HSP60] in patients with ischemic heart disease [IHD]. 62 patients with IHD having either acute myocardial infarction [AMI; n=31] or unstable angina [UA; n=31] and 31 sex- and age- matched healthy subjects as a control group were enrolled in this study. Serum samples of participants were tested for the presence of hs-CRP and antibodies against C. pneumoniae and Cp-HSP60 using ELISA method. The seroprevalence of anti-C. pneumoniae antibody in AMI group [93.5%] or UA group [90.3%] was significantly higher than the control group [61.3%; p<0.001]. The seroprevalence of anti-Cp-HSP60 IgG was 22.6% in healthy subjects with mean end titer of 43.1 +/- 6.32. The seropositive rates of anti-Cp-HSP60 were 48.4%, 54.8% and 51.6% in AMI, UA and the overall IHD groups with mean end titers of 94 +/- 22.86, 113.8 +/- 24.25 and 103.9 +/- 16.57, respectively. Both the seroprevalence and the mean titer of anti-Cp-HSP60 in patients groups were significantly higher than those observed in the control group [p<0.04 and p<0.03, respectively]. Moreover, the mean serum hs-CRP levels was significantly higher in the IHD group as compared to the control group [21.6 Mu g/ml +/- 3.73 vs 2.5 Mu g/ml +/- 0.52; p<0.00001]. The mean serum hs-CRP levels of AMI [30.3 Mu g/ml +/- 6.07] or UA [12.9 Mu g/ml +/- 3.85] groups were also significantly higher than those observed in the control group [p<0.00001 and p<0.001, respectively]. Furthermore, the difference of the mean serum hs-CRP levels between AMI and UA groups was also significant [p<0.02]. These results showed that the seroprevalence of antibodies against C. pneumoniae and Cp-HSP60 and the serum levels of hs-CRP and anti-Cp-HSP60 IgG were higher in patients with IHD