Small vessel occlusion and syphilis in patients with first-ever ischemic stroke

ABSTRACT Background: Syphilis is an endemic disease, particularly in low- and middle-income countries, with vascular involvement in large vessels (aortitis), but no clear relationship with stroke patients, except for those who presented with meningovascular neurosyphilis. Objective: To investigate the relationship between a positive history of syphilis determined by serological testing and ischemic stroke etiology, particularly small vessel disease (SVD). Methods: In total, 269 first-ever ischemic stroke patients admitted to the stroke unit were tested for syphilis. Patients with neurosyphilis were excluded. All patients were classified according to the ASCOD phenotyping as SVD — when SVD was the potential causal mechanism (S1) — or non-SVD — when SVD was uncertain (S2), unlike (S3), or not detected (S0). Results: Syphilis was positive in 32 (12%) patients. When comparing patients with positive and negative serology, the only significant difference was SVD as the causal mechanism (S1) in patients with positive results: 9 (28%) vs. 22 (9%), p<0.01. Conclusion: The current study showed that the frequency of positive syphilis serological test was higher in patients with first-ever ischemic stroke and SVD as the potential causal mechanism. This finding could be related to the endothelial dysfunction occurring in syphilis.


INTRODUCTION
Syphilis is a sexually transmitted systemic infection recognized since the late 14 th century 1,2 , caused by the Treponema pallidum pallidum 3 . Early and late forms have been described, with the latter further divided into latent (asymptomatic) and tertiary syphilis, which, in turn, can be classified as cardiovascular syphilis, neurosyphilis (NS), and peripheral gummatous syphilis. NS has a broad clinical spectrum, with five major subcategories: syphilitic meningitis, meningovascular NS, general paresis, tabes dorsalis, and ocular NS 4 .
Besides the relationship between NS and stroke, Kelley et al. were some of the first to call attention to the fact that stroke patients had a higher rate of positive syphilis serological tests compared to controls (24 versus 15%), although no correlation between stroke type and positive results was found 5 .
Today, we have a better understanding of the pathophysiology of both conditions, particularly endothelial dysfunction, as shown by studies of lesions from secondary syphilis 6 . This study aimed to determine the relationship between a positive history of syphilis determined by serological testing and first-ever ischemic stroke (FEIS), exploring its association with the mechanism of cerebral ischemia, especially small vessel disease (SVD).

METHODS
This is a retrospective analysis of a prospective database of patients with FEIS admitted to the Hospital de Clínicas of the Universidade Federal do Paraná from January 2012 to July 2017. The study was approved by the local Research Ethics Committee, and the board waived the need for patient consent.
All patients underwent etiological investigation with at least computed tomography (CT) scans (one at admission and the other 24 h after symptom onset), electrocardiography, transthoracic echocardiography, and vascular ultrasound. Other tests, such as magnetic resonance imaging (MRI), 24-hour Holter monitoring, cerebrospinal fluid (CSF) analysis, CT angiography, and angiography, were performed on a case-by-case basis. Syphilis testing was carried out in two steps, following the Brazilian Ministry of Health guidelines 7 : first, a treponemal test (chemiluminescence) and, if positive, a non-treponemal test (venereal disease research laboratory -VDRL); when the treponemal test was positive, but the VDRL was non-reactive, a second treponemal test (indirect immunofluorescence: fluorescent treponemal antibody absorption -FTA-Abs) was performed to exclude a false-positive chemiluminescence result. Tests were considered positive when both chemiluminescence and VDRL were positive or when chemiluminescence and FTA-Abs were concordant. Cases with only a positive VDRL (with negative treponemal test) were categorized into the non-syphilis group due to the lack of specificity of the test. Patients in the syphilis group were submitted to CSF analysis. Cases with a reactive CSF-VDRL, pleocytosis, or elevated protein were subcategorized in the NS group 7 , as presented in Figure 1.
The variables studied were age, sex, cerebrovascular risk factors (hypertension, diabetes, hyperlipidemia, and tobacco use), presence of atrial fibrillation, National Institutes of Health Stroke Scale/Score (NIHSS) at admission, modified Rankin Score (mRS) at discharge and at the last visit, stroke recurrence, and mortality. The stroke mechanism was classified according to the TOAST phenotype 8 in large-artery atherosclerosis, cardioembolism, small vessel occlusive disease, stroke of other determined etiology, and stroke of undetermined etiology. All patients were also evaluated based on the ASCOD phenotyping 9 and classified into two groups: SVD -when SVD was potentially causal (S1) -and non-SVD -when SVD was uncertain (S2), unlike (S3), or not detected (S0).
Statistical analysis was performed in the software Statistica 8.0; NIHSS was presented as median and first (Q1) and third (Q3) interquartile range (median [Q1-Q3]). We assessed the statistical significance using Student's t-test for continuous variables and Fisher's exact test or chisquare test for qualitative variables. Statistical significance was set at p<0.05, with a 0.95 confidence interval.

RESULTS
Out of the 273 FEIS patients tested for syphilis in the study period, four were diagnosed with NS. Among the 269 cases that fulfilled the study inclusion criteria, 32 (12%) had a positive syphilis serological test, while the other 237 (88%) had negative results. When comparing patient characteristics between these two groups, 10 (31%) individuals in the positive result group presented a higher frequency of small vessel occlusive disease based on TOAST, in contrast to only 35 (14%) in the negative result group, p=0.039. The ASCOD phenotyping analysis revealed nine (28%) patients in the syphilis group with SVD as a potential cause (S1) compared to 22 (9%) in the non-syphilis group, p=0.004. No differences were found in other stroke subtypes or between these groups, including those related to outcome. Table 1 presents the demographic data and results.

DISCUSSION
The current study showed a higher frequency of positive syphilis tests in patients with stroke, predominantly in those with SVD as the potential cause of the event.
Syphilis is considered an endemic disease in Latin America, with an estimated prevalence of positive tests of 2.6% in the general population 3,10 . This investigation identified 12% of positive tests in stroke patients, which is close to previous studies, whose values were between 4 and 8.4% 11,12 .
The association between syphilis and SVD observed in the present study could result from recent pathophysiological data. Some proteins expressed by Treponema pallidum, such as Tp0965 13 and Tp17 14 , have demonstrated the capacity to activate endothelial cells, ultimately leading to the up-regulation and expression of vascular endothelial growth factor (VEGF) and adhesion molecules by the endothelium, such as ICAM-1, E-selectin, and VCAM-1 13,14 . Furthermore, studies of primary and secondary syphilis lesions have documented the role of interleukin-6 (IL-6) and increased vascular permeability during their development 15 . All of these features are common to the pathogenesis of SVD, as recently described 16 . Therefore, an inflammatory endothelial mechanism could play an important role in a pathological process, eventually resulting in the development of SVD and stroke, a relationship that resembles the one associated with stroke in general 17 and myocardial infarction specifically 17,18 .
The clinical impact of the syphilis diagnosis could not be confirmed, as both groups had similar results to all other variables, including outcome. Therefore, even if syphilis could accelerate the endothelial dysfunction, this acceleration might depend on other stroke risk factors, which could even trigger or exacerbate a vascular inflammatory response and consequent endothelial dysfunction associated with SVD.
This study has limitations, such as the small sample, the retrospective analysis, and the assessment of only clinical features; no biomarker study was performed to confirm the hypothesis of this relationship, but the results replicate previous findings in medical literature and further raise the question of a common pathway for distinct pathologies.
In conclusion, more than 10% of FEIS patients presented positive syphilis serological tests, which were more common in the small vessel occlusive disease mechanism, based on two different classifications. This finding could be related to the endothelial dysfunction occurring in syphilis. Future studies could focus on biomarkers related to endothelial dysfunction and their relationship with SVD and syphilis.