The difference of lipid profiles between psoriasis with arthritis and psoriasis without arthritis and sex-specific downregulation of methotrexate on the apolipoprotein B/apolipoprotein A-1 ratio

Background Methotrexate (MTX) has a protective effect against cardiovascular diseases (CVD), but the mechanism is unclear. Objective To investigate the effect of MTX on lipid profiles and the difference between psoriasis without arthritis (PsO) and psoriatic arthritis (PsA). Methods In this prospective study, we recruited 288 psoriatic patients (136 PsA and 152 PsO) who completed 12 weeks of MTX treatment. Total cholesterol (TC), triglycerides (TG), lipoprotein A [LP(a)], high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein (LDL), apolipoprotein A1 (ApoA1), and ApoB were measured. Results Compared with sex- and age-matched healthy controls, psoriatic patients had significantly (p < 0.0001) higher levels of proatherogenic lipids and lower levels of anti-atherogenic lipids. PsA patients had a higher ApoB/ApoA1 ratio than PsO patients (p < 0.05). Stepwise regression analysis found a positive correlation between the inflammatory marker hCRP and the Psoriasis Area Severity Index (PASI), ApoB/ApoA1 ratio, BMI, and smoking. ApoB was positively associated with concomitant arthritis, diabetes, and hypertension. MTX decreased the levels of pro-atherogenic and anti-atherogenic lipids. However, a significant reduction of the ApoB/ApoA1 ratio by MTX was only observed in male patients. Conclusion PsA patients had a significantly higher percentage of concomitant disease than PsO. The decrease of MTX on CVD might be related with sex. Trial registration ChiCTR2000036192


Introduction
Psoriasis is a common chronic inflammatory disease characterized by keratinocyte abnormalities and immune dysfunctions. It affects about 2-3% of the world population [1]. About 5.8 to 30% psoriatic patients without arthritis (PsO) will develop psoriatic arthritis (PsA) [2,3]. Epidemiological and clinical studies have consistently shown that psoriasis is associated with an increased cardiovascular risk [4]. The inflammatory cytokines found in psoriatic lesions may cause insulin resistance and trigger endothelial cell dysfunction leading to atherosclerosis and ultimately resulting in stroke or myocardial infarction [5]. Compelling evidence further suggests that proinflammatory cytokines such as interleukin (IL)-1, IL-6, and tumor necrosis factor (TNF)-α may alter the function of hepatocytes and arterial smooth muscular cells to induce alternated lipoprotein compositions, enhance expression of cellular adhesion molecules, and increase lipid deposition on arterial walls. All of these features can contribute to the development of arterial plaques [6]. Cytokines may then destabilize the plaque by promoting the rupture of fragile neo-vessels and increasing the expression of the plaque's fibrous cap. This cascade of events may ultimately lead to plaque rupture and the formation of life-threatening thrombi [7].
Psoriasis is significantly associated with a higher prevalence and incidence of dyslipidemia [8]-a known risk factor for cardiovascular disease. Dyslipidemia is a broad term that describes any abnormality of plasma lipids including perturbations in plasma lipid levels or abnormalities in lipid composition. Multiple measurements of dyslipidemia are significantly affected and include raised triglycerides, raised LDL cholesterol, lowered HDL cholesterol, raised cholesterol, and raised lipoproteins [9][10][11]. Due to the remarkable heterogeneity of defining dyslipidemia, there is a lack of sex-and age-matched healthy controls. Moreover, adjustments were not made for confounding factors in most studies. Thus, the association between psoriasis and dyslipidemia still remains unclear.
Arthritis is an important determinant for psoriatic patients to develop severe vascular events in Taiwan [12]. Moreover, some arthritic patients on low-dose methotrexate (MTX) have altered blood lipids versus those not taking MTX [13]. In this prospective, crosssectional study, we compared the differences in lipid profiles and cardiovascular risk parameters between PsA patients and sex-and age-matched healthy controls, between PsO patients and sex-and age-matched healthy controls, and between male and female psoriatic patients. Furthermore, we analyzed the effect of MTX on lipid profiles and further analyzed the influencing factors on lipid profiles and cardiovascular risk parameters after adjustments for sex, age, age at disease duration, disease duration, Psoriasis Area Severity Index (PASI), body mass area (BSA) scores at baseline, smoking, alcohol, height, weight, body mass index (BMI), hypertension, and diabetes.

Patients
This single-center prospective trial was performed in the Department of Dermatology, Huashan Hospital, Fudan University, between December 2, 2015, and December 2, 2019. In total, 288 psoriatic patients who received oral MTX treatment for 12 weeks and 288 sex-and age-matched healthy controls without prior medication from a medical examination center were recruited. The medical ethics committee of Huashan Hospital at Fudan University reviewed and approved the protocol (approval #MTX201501); all patients provided written informed consent. Patients aged ≥18 years were recruited from the outpatient population. The diagnosis of psoriasis and psoriatic arthritis (n = 136) was based on typical clinical and/or histopathological criteria and the Classification Criteria for Psoriatic Arthritis (CASPAR classification criteria), respectively. Patients who received systemic treatments (acitretin, cyclosporin, glucocorticoids) for arthritis or psoriasis at 1 month were excluded. The topical treatments had been stopped for more than 1 week before the beginning of the study. The therapeutic regimen followed the European guidelines on contraindications and restrictions on methotrexate. None of the patients used lipidlowering drugs.

Treatment
The initial oral MTX dose was 7.5-10 mg once weekly. The dose was increased by 2.5 mg every 2 to 4 weeks to a maximum of 15 mg weekly depending on the patient's clinical response, side effects, and hematology/chemistry tests. If liver enzyme elevations were >2-and <3-fold, then the MTX dose was reduced by 2.5 mg weekly and administered once 2-4 weeks later. MTX treatment was stopped if the liver enzyme elevations were >3-fold [14].

Assessments of lipid profiles and disease characteristics
Two certified dermatologists graded the severity and extent of psoriasis using the Psoriasis Area Severity Index (PASI) and body surface area (BSA) scores. Lipid profiles at baseline and 12 weeks for MTX treatment and fasting blood glucose at baseline were measured using conventional laboratory techniques at Huashan Hospital. Sex, age, age at disease onset, smoking, alcohol, hypertension, diabetes, height, weight, and body mass index (BMI) were recorded.

Statistical analysis
Data are expressed as the means ± standard deviations (SDs). Statistical analyses were performed using the Mann-Whitney test, unpaired t-test, unpaired t-test with Welch correction, paired t test, χ [2] test, or Fisher's exact test as appropriate. Stepwise multiple regression analysis was performed after adjustments for sex, age, age at disease onset, disease duration, height, weight, body mass index (BMI), hypertension, diabetes, smoking, alcohol consumption, and PASI/BSA scores at baseline. Data analyses were performed using Graph Pad Prism version 5 (Graph Pad Software Inc) and SPSS ver. 23.0 software (SPSS Inc., Chicago, IL, USA). A p-value of <.05 (or p < .025 after multiple test correction) was considered to be statistically significant.

Results
The improvement of skin lesions in psoriasis without arthritis (PsO) was superior to that of psoriasis with arthritis (PsA) by methotrexate had a significantly older age (p < .0001), age at disease onset (p = .0052), longer disease duration (p = .0061), and higher percentage of hypercholesterolemia (p = .0087) than PsO patients. The mean PASI score at 12 W in PsO patients (p = .0354) was significantly lower than that in PsA patients although there was no difference in PASI score at baseline. The mean PASI (p = .0236) and BSA (p = .0105) change from baseline was significantly higher in PsO patients than that in PsA patients. In addition, the percentage of hypertension in PsA patients was significantly higher than that in PsO patients (p = .0029).

The association between the inflammatory marker hCRP, lipid profiles, and cardiovascular risk parameters in psoriatic patients
As shown in Table 5, stepwise regression analysis demonstrated that serum hCRP level was positively correlated with PASI score at baseline (p=.000), ApoB/ ApoA1 ratio (p=.011), BMI (p=.003), and smoking (p=.015). Serum ApoA1 level was positively related with age (p=.006) and negatively associated with sex (p=.015), weight (p=.004), and hCRP (p=.006). Serum HDL level was negatively correlated with sex  (p=.000) and weight (p=.014). Serum ApoB level was positively associated with diastolic blood pressure (p=.044), the concomitant of arthritis (p=.001), and diabetes (p=.006). Serum TC level was negatively related with sex (p=.031) and positively correlated with age (p=.000), diastolic blood pressure (p=.015), and the concomitant of arthritis (p=.002). Serum LDL level was positively associated with diastolic blood pressure (p=.003) and age at disease onset (p=.014).

Discussion
Our results demonstrate that PsA and PsO patients have significantly higher levels of pro-atherogenic lipids such as ApoB and LDL and lower levels of antiatherogenic lipids such as ApoA1 than sex-and agematched healthy controls. The ratios of ApoB/ApoA1 and LDL/HDL-C were also significantly higher in PsA and PsO patients than in healthy controls. The ApoB/ ApoA1 ratio is a strong and new risk factor for cardiovascular disease (CVD) [15]. Data from Indian patients with acute myocardial infarction (AMI) demonstrated that the ApoB/ApoA1 ratio was a better discriminator of coronary artery disease (CAD) risk than other conventional lipid ratios including the ratio of TC/ HDL-C and LDL/HDL-C [16]. Moreover, the ApoB/ ApoA1 ratio was associated with femoral artery atherosclerosis as measured both as intima-media thickness and plaque occurrence [17]; and carotid artery atherosclerosis measured by intima-media thickness [18]. A higher ApoB/ApoA ratio implies that more cholesterol is likely to be deposited in the arterial wall thereby provoking atherogenesis [19]. Our results are in accordance with a previous observation that psoriatic patients had a higher prevalence of ischemic heart disease (3.3% vs 1.8%, OR 1.87) and vascular cerebral accidents (1.8% vs 1.2%, OR 1.55) [20]. We report a significantly higher percentage of hypertension and hypercholesterolemia in PsA patients than PsO patients probably because the PsA cohort was older at disease onset with longer disease duration [21]. Previous studies demonstrated that psoriatic patients have a greater prevalence and incidence of hypertension. In particular, PsA and severe psoriasis were associated with a greater odds of hypertension [22]. Psoriatic patients also have increased renin-angiotensin system activity, vascular damage, and oxidative stress linked to psoriasis and hypertension [23][24][25].
In addition, we found that PsA patients had significantly higher levels of ApoB, TC, and LDL and a higher ApoB/ApoA1 ratio than PsO patients, which is in accordance with a previous report that PsA patients had higher burdens of specific comorbid disease than PsO patients [26]. Serum ApoB and TC levels were positively associated with arthritis after adjusting for sex, age, weight, BMI, disease duration, PASI and BSA score, diabetes, and hypertension. Our results on a higher ApoB/ApoA1 ratio in PsA differ from a recent study, which reported a higher ApoA/ApoB ratio in PsA compared to PsO [27]. Discrepancy in the study population might explain these results, since only age-but not sex-matched healthy controls were investigated in the former study. Indeed, our study found sex was also an important factor influencing lipid profiles. Male patients had significantly lower levels of TC, especially anti-atherogenic lipid profiles such as high-density lipoprotein HDL-C and ApoA1 than female patients. The ratio of ApoB/ApoA1 was significantly higher in male patients than in female patients, which was consistent with the epidemiology of cardiac and vascular disease whereby the age-adjusted CVD mortality and morbidity rates are highest in men than in women [28].
The effects of MTX on lipid profiles and CVD are controversial. One meta-analysis showed that the use of MTX predicted a reduction of 20% in the risk of cardiovascular events [29]. However, another study found that there was no effect of MTX on the levels of HDL, LDL, TG, TC, IL-1β, IL-6, and CRP. Low-dose MTX did not result in fewer cardiovascular events among patients with stable atherosclerosis than placebo. But this study did not stratify the patients by sex [30]. In our study, when patients were stratified by PsO and PsA, MTX significantly downregulated the levels of serum ApoB, TC, HDL-C, and Lp(a), as well as the ratios of ApoB/ApoA1 and LDL/ApoB in patients with PsO and PsA. Furthermore, MTX differentially reduced the levels of serum ApoA1 and LDL in patients with PsO and decreased serum TG level in patients with PsA. When patients were stratified by sex, MTX significantly downregulated the levels of serum ApoB, TC, and Lp(a) in both male and female patients with psoriasis, and differentially reduced the levels of serum ApoA1, HDL-C, LDL, and ratios of ApoB/ApoA1 and LDL/ApoB in male patients with psoriasis, but not in female patients. Therefore, the decrease of ApoB, TC, and Lp(a) by MTX was not related with sex and psoriasis subtype. However, the effect of MTX on some lipid profiles (ApoA1, HDL-C, LDL, TG) was associated with sex and psoriasis subtype. The mechanism remains to be clarified. Our another study also found MTX treatment significantly reduced the levels of serum hCRP and blood pressure in male patients, not in female patients. This indicates that the downregulation of MTX on lipid profiles was related with anti-inflammation. It has been reported that androgens can enhance the anti-inflammatory effects of MTX, which might be the main reason why the antiinflammatory effect and downregulation of lipid profiles of MTX were more obvious in male patients than in female patients [31].
The association between inflammation and lipid profiles was further analyzed in this study. Our results demonstrated that the inflammatory biomarker hCRP was not only positively correlated with disease severity (PASI score) at baseline, but was also positively associated with ApoB/ApoA1. In addition, high BMI and smoking are known to increase serum hCRP level. This might be one reason why men had significantly lower levels of anti-atherogenic lipid profiles. The ratios of ApoB/ApoA1, TC/ HDL, LDL/HDL-C, and LDL/ApoB were all regarded as cardiovascular risk parameters. But only ApoB/ApoA1 was positively correlated with inflammatory marker hCRP and the concomitant of arthritis, and they were all associated with diastolic blood pressure or the concomitant of hypertension, weight, or BMI in a stepwise regression analysis. In addition, TC/HDL-C ratio was positively related with the concomitant of diabetes. Therefore, ApoB/ApoA1 ratio is the best biomarker to predict the risk of CVD [32].
In addition, we further analyzed the association of lipid profiles with clinic characteristics of psoriasis and concomitant disease. Our data showed that ApoA1 level was negatively associated with hCRP, not HDL level. This indicates that ApoA1 is more sensitive to inflammation than HDL-C. Another study also found that serum ApoA1 level showed a strong negative correlation with some markers of systemic inflammation including serum CRP and interleukin (IL)-8 levels and blood neutrophil count [33]. ApoB is the best marker to predict concomitant disease, which was positively correlated with diastolic blood pressure, the concomitant of diabetes and arthritis. ApoB is a key component of atherogenic lipids and a more accurate measure of cardiovascular risk than LDL or non-HDL-C. ApoB was reported to aggravate arthritis by eliciting the production of TNF-α, IL-1β, and IL-6 through p38 mitogenactivated protein kinase and NF-κB pathways. ApoB is an important independent immune-modulator that links lipid metabolism with local and systemic inflammatory responses [34]. TC and LDL were positively associated with diastolic blood pressure, and TG was positively correlated with diabetes. It has been reported that TC might correlate to arterial stiffness [35] and increase the lifetime risk of coronary heart disease mortality [36]. Diabetes was reported to reduce HDL cholesterol, a predominance of small dense LDL particles, and elevated triglyceride levels [37]. Therefore, the levels of lipid profiles were related with psoriasis and its concomitant disease.

Conclusions
In conclusion, inflammation in psoriasis can increase serum hCRP levels, resulting in the reduction of antiatherogenic lipid ApoA1 and increase of pro-atherogenic lipid ApoB. ApoB and ApoB/ApoA1 ratio were the best biomarkers to predict inflammation and concomitant disease, especially CVD. MTX decreased pro-atherogenic lipid profiles as well as anti-atherogenic lipid profiles. The ratio of ApoB to ApoA1 was significantly downregulated in male patients with psoriasis after MTX treatment, indicating that the role of MTX on CVD and other concomitant diseases might be related to sex.