Objective To characterise associations between individual nailfold capillary aberrations with autoantibodies in a cross-sectional study on children and adults with Raynaud’s phenomenon (RP).
Methods Consecutive children and adults with RP and without previously known connective tissue disease (CTD) systemically underwent nailfold capillaroscopy and laboratory tests for the presence of antinuclear antibodies (ANA). The prevalence of individual nailfold capillary aberrations and ANA was assessed, and the associations between individual nailfold capillary aberrations and ANA were analysed separately in children and adolescents.
Results In total, 113 children (median age 15 years) and 2858 adults (median age 48 years) with RP and without previously known CTD were assessed. At least one nailfold capillary aberration was detected in 72 (64%) of included children and in 2154 (75%) of included adults with RP (children vs adults p<0.05). An ANA titre ≥1:80, ≥1:160 or≥1:320 was observed in 29%, 21% or 16% of included children, and in 37%, 27% or 24% of screened adults, respectively. While the occurrence of individual nailfold capillary aberrations was related to the presence of an ANA titre of ≥1:80 in adults (reduced capillary density, avascular fields, haemorrhages, oedema, ramifications, dilations and giant capillaries: each p<0.001), no comparable association between nailfold capillary aberrations and ANA was observed in children with RP without previously known CTD.
Conclusion In contrast to adults, the association between nailfold capillary aberrations and ANA might be less pronounced in children. Further studies are warranted to validate these observations in children with RP.
- Autoimmune Diseases
- Cardiovascular Diseases
Data availability statement
Data are available upon reasonable request.
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WHAT IS ALREADY KNOWN ON THIS TOPIC
Nailfold capillaroscopy and laboratory tests comprising antinuclear antibodies (ANA) are routinely obtained in parallel in patients with incipient Raynaud’s phenomenon (RP) to screen for an underlying connective tissue disease (CTD).
WHAT THIS STUDY ADDS
The present study examined relations between nailfold capillary pathologies and elevated ANA-titres in both children and adults with incipient RP. Compared with adults, the relation of nailfold capillary pathologies and elevated ANA-titres appeared less pronounced in children with incipient RP.
HOW THIS STUDY MIGHT AFFECT RESEARCH, PRACTICE OR POLICY
This study suggests that in children with incipient RP, screening examinations for and underlying CTD should involve nailfold capillaroscopy and ANA, since there appears to be no clear association between both markers.
Raynaud’s phenomenon (RP) is a common vasospastic disorder that potentially causes paroxysmal ischaemic attacks, predominantly affecting fingers and toes. In adolescence, RP affects up to 15% above the age of 15 years.1 Focusing on young patients, RP most commonly manifests after the age of 10 and more frequently affects women than men.2
RP can be classified as primary RP of unknown cause or secondary RP resulting from an emerging local or systemic disease.3 The diagnostic workup of patients with incipient RP primarily comprises the screening for an underlying connective tissue disease (CTD).4 5 This screening process comprises nailfold capillaroscopy (NC) and laboratory tests for immunological markers, such as antinuclear antibodies (ANA) and respective subsets.5–10 The predictive values of NC and immunological markers are well established in adult patients with RP.11 12
By analogy, in children, NC and serological tests for ANA are commonly applied when assessing incipient RP.13–16 However, the predictive value of NC in children with RP is less clear. Compared with adults, epidemiological data suggest a lower specificity of individual nailfold capillary changes in children with RP.17 18 While specific capillary aberrations are related to elevated levels of ANA in adults with RP, the relation of specific nailfold capillary changes to increased titres of ANA in children is unknown.19
We, therefore, aimed to assess the relation of specific NC aberrations to increased ANA-titres in children with RP and without previously known CTD and compare these observations to adult patients with RP.
Consecutive patients with incipient RP and without previously known CTD, who were admitted to the Division of Angiology outpatient clinic at the Medical University of Vienna, were included in this analysis. A detailed description of the study collective was published previously.12 In brief, the diagnosis of RP was established according to the clinical presentation by a senior vascular physician.20 21 After excluding macrovascular obstructions by standardised pulse volume measurements, all patients with RP (and without previously known CTD) were referred to NC and serological testing for ANA. For the present analysis, patients aged <18 years were categorised as children, while patients aged ≥18 years were classified as adults.
NC was performed under standardised conditions in a quiet room at an ambient temperature of 23.3°C±0.6°C. For acclimatisation, patients had to rest in a sitting position for 20 min before NC. All study participants were advised to refrain from manicure at least 1 week prior to NC and remove any nail polish ahead of the examination. A CAM1 capillaroscopic system (KK Research Technology LTD and Leitz, Wetzlar) was used for all examinations at 250/300-fold magnification. In each participant, the fingers 2 to 5 of both hands were evaluated, and nailfold capillaries were classified into a regular nailfold capillary pattern when no nailfold capillary aberration was found, or an irregular nailfold capillary pattern when at least one of the following capillary aberrations was detected: (1) a reduced capillary density (<30 hairpin-shaped capillaries per 5 mm nailfold), (2) avascular fields (large capillary-free area), (3) dilated capillaries (maximum capillary diameter >20 µm but ≤50 µm), (4) giant capillaries (maximum capillary diameter >50 µm), (5) haemorrhages (hemosiderin extravasations), (6) tortuous capillaries, (7) ramifications and (8) capillary oedema (capillary blurring or pericapillary halo).12 19 22 Representative images of individual capillary aberrations were published previously.12 Recorded images were processed by a computer equipped with the corresponding software (CapiScope, KK technology). All examinations were supervised by an experienced investigator (MEG), and two investigators finally evaluated images in consensus (OS and MEG).
Venous blood samples were drawn from all patients, and ANA titres were determined using indirect immunofluorescence on Hep-2 cells (Kallestad, Bio-Rad Laboratories, Hercules, California) at the institutional laboratory. For further analyses, patients were categorised into the following groups of ANA titres: ≥1:80, ≥1:160 and ≥1:320. Since these titres refer to laboratory dilution series, patients with an ANA titre ≥1:160 were allocated to both groups, ANA ≥1:80 and ANA ≥1:160, for the analysis of associations with NC. Similarly, patients with an ANA titre ≥1:320 were allocated to all three groups, ANA ≥1:80, ANA ≥1:160 and ANA ≥1:320, for assessing the associations with NC.
In case of specific immunofluorescence patterns on Hep-2 cells or clinically suspected CTD, ENA-subsets (anti-Scl-70, anti-CENP-B, anti-SSA (Ro), anti-SSB (La), anti-U1-RNP, anti-Sm, anti-Jo-1 and anti-ds-DNA antibodies) were determined.23 While ENA-subsets were determined by Ouchterlony immunodiffusion (Auto I.D., Immuno Concepts, Sacramento, California) from 1994 to September 2005, multiplex fluorescent technology (FIDIS Connective 10, BMD, France) was used between September 2005 and January 2007, and UniCAP Elia System (Phadia, Sweden) hereafter and until the end of the survey period.
Discrete variables are given as counts and percentages, and continuous variables are presented as the median and IQR. Group characteristics were compared using Fisher’s exact tests, χ² tests and Mann-Whitney-U tests as appropriate. A logistic regression model was used to calculate the ORs and 95% CIs of irregular nailfold capillaries, defined as the presence of at least one capillary aberration, for ANA-titres ≥1:80, ≥1:160 and≥1:320, respectively. In addition, the OR and 95% CI of individual nailfold capillary aberrations (reduced capillary density, avascular fields, haemorrhages, capillary oedema, ramifications, dilations, giant capillaries, tortuous capillaries) for ANA-titres ≥1:80, ≥1:160 and ≥1:320 were analysed, respectively.
All statistical calculations were done using the software package R V.3.6.1 (R Core Team (2020). R Foundation for Statistical Computing, Vienna, Austria. URL https://www.R-project.org/).
A total of 2971 consecutive patients with RP and without previously known CTD were included in this analysis. Of these patients, 113 were children (87 women, 77%) with a median age of 15 years (IQR 14–17 years), and 2858 were adults (2222 women, 78%) with a median age of 48 years (IQR 36–59 years). Smoking status was positive in 3 (11%) of children and in 246 (35%) of adults.
Nailfold capillary aberrations in children and adults
Irregular nailfold capillaries (presence of ≥1 individual nailfold capillary aberration) were detected in 72 (64%) children and in 2154 (75%) adults with RP (children vs adults p<0.05). Within the spectrum of individual nailfold capillary aberrations, tortuous capillaries were less frequently observed in children than in adults. The prevalence of other nailfold capillary aberrations (reduced capillary density, avascular fields, haemorrhages, capillary oedema, ramifications, dilations, giant capillaries) did not differ between children and adults (table 1).
Autoantibodies in children and adults
An ANA-titre of ≥1:80 was observed in 33 (29%) children and 1059 (37%) adults with RP (children vs adults p<0.09). An ANA-titre of ≥1:160 was found in 24 (21%) children and 768 (27%) adults (children vs adults p=0.14) and an ANA-titre of ≥1:320 was detected in 18 (16%) children and 671 (23%) adults (children vs adults p=0.06). The percentage proportions of respective ANA subsets in children and adults with RP are depicted in table 2.
Relation between capillary aberrations and autoantibodies
In children, we found no association between the presence of irregular nailfold capillaries (presence of ≥1 individual nailfold capillary aberration) and ANA-titres ≥1:80 (OR 0.78, 95% CI 0.34 to 1.81, p=0.57), ≥1:160 (OR 1.03, 95% CI 0.40 to 2.70, p=0.95) or ≥1:320 (OR 0.84, 95% CI 0.30 to 2.36, p=0.74). In adults, the presence of irregular nailfold capillaries was related to ANA-titres ≥1:80 (OR 1.68, 95% CI 1.39 to 2.02, p<0.001), ≥1:160 (OR 1.83, 95% CI 1.48 to 2.27, p<0.001) and ≥1:320 (OR 2.24, 95% CI 1.77 to 2.84, p<0.001).
The percentage proportions of positive ANA-titres (1≥1:80, 1≥1:160 and 1≥1:320) within subgroups of individual nailfold capillary aberrations are shown in figure 1.
In children, none of the individual nailfold capillary aberrations—except tortuous capillaries—was associated with increased levels of ANA. In adults, however, every single nailfold capillary aberration—except tortuous capillaries—was related to elevated ANA-titrers (figure 2, table 3).
The main finding of this study is the discrepancy in the associations of irregular nailfold capillaries with ANA between children and adults. While most nailfold capillary aberrations—except tortuous capillaries—were related to elevated ANA-titres in adults with RP, no comparable association between individual nailfold capillary aberrations and ANA was found in children with RP.
These observations need to be interpreted considering the assessed study population: both groups, children and adults, consisted of allegedly healthy subjects who presented with incipient RP without a previously known CTD. Regarding the demographic characteristics of included children, the median age is consistent with the age distribution of previous studies in children with RP.1 Furthermore, the female predominance in the study population is typical for RP and in line with earlier publications on paediatric patients with RP.2 The study population, therefore, reflects a real-world setting in specialised outpatient clinics performing screening examinations in patients with RP.
Referring to CTD screening in RP, the underlying objective of the present study warrants clarification: since CTD screening in RP commonly comprises NC and serological tests for ANA in parallel, we were interested in whether NC could serve as a prescreening tool in childhood to anticipate the need for blood draws. While, in our study, the overall prevalence of irregular nailfold capillaries was lower in children than in adults, the percentage distribution of individual nailfold capillary aberrations and their prevalence were similar in children and adults with RP. This could be interpreted as a resemblance in the development of microangiopathy in children and adults with incipient RP.
Furthermore, the prevalence of ANA and ANA subsets appeared to be similar in children and adults who underwent CTD screening because of incipient RP.
To analyse these observations, however, the following considerations warrant mention: first, neither the presence of ANA nor the presence of irregular nailfold capillaries by themselves can sufficiently establish the diagnosis of a CTD. In particular, capillary aberrations can be regarded as a potential indicator for the development of microangiopathy, as can be observed in the scope of several CTD. Especially the progression of RP to systemic sclerosis might be predicted by a typical course of nailfold capillary abnormalities.8 11 24 Second, most patients presenting with incipient RP can be classified as primary RP since an underlying systemic disease can only be detected in 11% of (adult) Raynaud patients.25 Finally, the association between pathognomonic NC aberrations and the occurrence of a CTD might be less pronounced in children than in adult patients with RP.17
Accordingly, the lack of association between capillary pathologies and ANA-positivity in children may be explained by the fact that the development of CTD-associated microangiopathy is a dynamic process with early capillary changes being subtle and probably less pronounced in children as compared with later disease stages in adult patients.8 26 Moreover, previous studies suggested a low prevalence of typical nailfold capillary aberrations in children with recently diagnosed juvenile rheumatoid arthritis, systemic lupus erythematosus or undifferentiated CTD.17 26 Especially in systemic lupus erythematosus, ANA-positivity may precede the diagnosis by years, and a state of ‘pre-autoimmunity’ may only apply to a subset of ANA-positive patients.27 Additionally, in juvenile idiopathic arthritis and paediatric dermatomyositis, the overall seroprevalence of ANA is <50%.28 29 The immunologic profile is also less characteristic in juvenile scleroderma, with only 77% of patients exhibiting positive ANA in a multicentre study including 153 participants.30 Both aspects could explain a ‘disjunction’ between microangiopathy and ANA-positivity in children with RP. Interestingly, we found no difference in the prevalence of giant capillaries in adolescent and adult participants. This capillary pathology has been reported as pathognomonic for CTD and practically absent in healthy individuals.22 In this respect, it should be noted that the regular nailfold capillary pattern in children resembles that observed in adults.31 Younger age, however, is associated with fewer capillaries, which may partially explain our findings.32 A recent study by Melsens et al, including patients with juvenile rheumatic and musculoskeletal diseases from 13 centres, reported capillary haemorrhages in 39.2% of matched healthy controls.26 In our study, capillary haemorrhages were the second most frequent capillary pathology, after dilations, in children with RP.
This study must be interpreted in light of its strengths and limitations. It should be acknowledged that the actual proportion of children with eventual secondary RP in this study population remains unknown since no clinical follow-up data were available. The proportion of children with secondary RP might have been lower than that of adult participants, considering the age distribution of most autoimmune rheumatic diseases.33 34 According to the cross-sectional character of NC assessments in this study, we cannot provide data on changes of capillary morphology over time. Future prospective studies with repetitive NC examinations are needed to assess NC changes over time. Neither was capillary morphology assessed in the standardised manner reported by Smith et al and we did not further distinguish degrees of reduced capillary density as suggested by Cutolo et al.35 Additionally, the smaller number of children compared with adults is a limitation. Nevertheless, the large overall size of the study population with available data from systematic NC and autoantibody screening examinations is a major strength and warrants mention. To validate the findings of the present investigation, prospective studies in children with incipient RP are needed.
The relation of nailfold capillary aberrations and the occurrence of ANA might be less pronounced in children than in adults. Additional studies are warranted to validate the findings of the present investigation.
Data availability statement
Data are available upon reasonable request.
Patient consent for publication
This study was approved by the institutional ethical committee (731/2010) and conducted in accordance with good clinical practice guidelines.
Figure 2 has been designed using Canva.com and assets from Freepik.com.
Contributors MM and OS conceptualised the study and drafted the manuscript. RR and MM were responsible for the statistical analysis. SS contributed to the creation of figures and tables. All authors provided critical feedback on the manuscript and approved the final version. s. All authors revised critically this manuscript and agree with its content. OS is responsible for the overll content of the paper.
Funding The authors have not declared a specific grant for this research from any funding agency in the public, commercial or not-for-profit sectors.
Competing interests None declared.
Provenance and peer review Not commissioned; externally peer reviewed.