This study was funded by a grant from Sun Health Technologies. BJR is the chief scientific officer for Sun Health Technologies, and the University of Illinois created a conflict management plan for minimizing bias and undue influence in running the trial and interpreting the data. This was necessary as BJR was the study neurologist. The funding source did not play any role in the writing of the manuscript and the decision to submit the manuscript for publication.
There is substantial evidence, from well-conducted epidemiological studies, that low vitamin D levels are correlated with increased risk for MS, and multiple case control studies have implicated the involvement of vitamin D deficiency in MS etiology. Narrow-band Ultraviolet B (NB-UVB; 300nm - 311 nm) induced vitamin D production has not previously been studied in a multiple sclerosis (MS) randomized placebo-controlled trial (RCT).
To investigate NB-UVB induced vitamin D production, immunomodulation and MS symptomology following NB-UVB phototherapy in a MS cohort.
Using a blinded RCT study design, twelve individuals 18 years or older with MS were enrolled and assigned (1:1) into individualized NB-UVB dose (10-30kJ/m) phototherapy, or into placebo treatment, delivered 3 times per week, for 8-weeks. Serum vitamin D levels, walking performance, strength, cognitive function, mood and circulating progenitor cells (CPCs: CD34+CD45dim), monocyte populations (Intermediate CD14+CD16+, Classical CD14+CD16-), and T regulatory cell (CD4+/CD25+/FoxP3+Tregs) count were assesed. The data were analyzed by 2 x 3 mixed factor ANOVA.
A statistically significant condition by time interaction on vitamin D levels (
UVB phototherapy corrects vitamin D deficiency. This study adds to the growing research investigating UVB treatment in MS.
Multiple sclerosis (MS) is an incurable debilitating autoimmune disorder. It is the most common nontraumatic chronic degenerative disease of the central nervous system (CNS). The worldwide MS prevalence, approximately 2.4 million people, follows an established geographic gradient that directly correlates with latitude and decreased sunlight intensity.
Vitamin D is a series of secosteroids obtained primarily through skin exposure to ultraviolet B radiation (UVB, 280-315 nm) from sunlight, and through both animal and vegetable dietary sources. The circulating 25(OH)D form of the hormone represents the principle vitamin D reservoir and is used as an indicator of overall vitamin D status. The primary approach for changing 25(OH)D levels in MS has involved supplementation with standard over-the-counter vitamin D preparations.
Sunlight may suppress MS through a 25(OH)D independent mechanism as well. More recent investigations into the inverse relationship between MS incidence and sunlight exposure led to the discovery that a narrow band of light (300–315 nm) prevents experimental autoimmune encephalomyelitis (EAE), which is the animal model of MS, without a change in 25(OH)D levels.
The purpose of this study is to evaluate the effects of UVB phototherapy (UVB lamp radiation; 300nm - 311 nm) on serum 25(OH)D levels, immune modulation and clinical benefit. We sought to determine whether UVB phototherapy increases 25(OH)D levels in individuals with MS, and we aimed to investigate whether UVB phototherapy effects circulating immunomodulatory cells or MS symptomology. We chose to investigate immunomodulatory cells, because MS is described broadly as being mediated by activated T-cell and B-cell lymphocytes, and because CNS inflammation is induced through myelin reactive T cells from the peripheral immune system.
Current MS therapies include targeting the destruction, depletion, or suppression of these autoreactive circulating T and B cell lymphocyte populations.
We performed a single center, triple-blind, randomized placebo-controlled, pilot trial. The study was registered with ClinicalTrials.gov, NCT02365259. The study protocol and ethics were approved by the University of Illinois at Urbana-Champaign (UIUC) Institutional Review Board (IRB). All participants provided written informed consent. Participant screening was undertaken over the telephone by the project coordinator, and participants were not enrolled until after receiving verification of inclusion and exclusion criteria status, and approval for participation by the study neurologist.
Community-residing women and men 18 years of age or older, with relapsing MS, who did not have any of the following exclusion criteria: (a) inability to stand still in a stabilized fashion without risk of falling for up to 6 minutes; (b) medical history of calcium disorder or knowledge of high calcium levels; (c) medical history of hyperparathyroidism; (d) current supplementation with oral vitamin D; (e) allergy to vitamin D; (f) history of cancer of any type including but not limited to skin cancer; (g) obesity defined as body mass index > 30; (h) history of fat malabsorption conditions (i.e., steatorrhea); (i) current use of anti-epileptic medication; (j) current use of glucocorticoids; (k) Fitzpatrick skin types I and VI; and (l) current use of tanning facilities, were recruited into the study. The study subjects were diagnosed with relapsing-remitting MS (RRMS) according to the McDonald criteria.
One randomization event occurred during the study. Study enrollees underwent baseline testing, and were randomized into an intervention condition (UVB phototherapy or Placebo control) by a computer-generated random sequence using standard methods. The final sample completing the study included 12 persons with MS who were evenly distributed between treatment conditions. The study participants, site personnel and clinical neurologist involved in the study were masked to the results of randomization. The University of Illinois created a conflict management plan for monitoring and minimizing bias and undue influence in running the trial and interpreting the data. The study phototherapy technician responsible for administering the intervention treatment was the only person aware of the intervention assignments.
Serum 25(OH)D level, circulating progenitor, monocyte subsets, andTreg numerical status, walking performance, hand-grip strength, cognitive function and mood were assessed at baseline, 4 weeks and again at 8 weeks after beginning the UVB phototherapy or placebo-controlled intervention. The UVB intervention condition consisted of an individualized regimen. Each sub-minimal erythema dose (10-30kJ/m2) of UVB (Sun Health FS UVB 298-313 nm, 40WT10) phototherapy delivered was based on skin sensitivity, and took into consideration subject Fitzpatrick skin type classification per the 2014 guidelines issued by the American Academy of Dermatology (AAD).
Venous blood was collected from an antecubital vein after an overnight fast at the same time of day for all assessments to quantify 25(OH)D levels. Blood was allowed to clot in serum tubes (BD Vacutainer: Franklin Lakes, NJ) for 60 min at room temperature, then serum was separated by centrifugation at 1300 × g for 15 minutes. Serum was aliquoted and stored at −80OC until analysis. 25(OH)D concentrations were quantified in serum by a standard protocol, and based on the manufacturer’s instructions (Immunodiagnostic Systems, United Kingdom). All samples were analyzed in triplicate, and 25(OH)D concentrations were expressed as nmol/L.
Circulating progenitor cells (CPCs: CD34+CD45dim), monocyte populations (Intermediate CD14+CD16−, Classical CD14+CD16+), and Tregs (CD4+/CD25+/FoxP3+Tregs) were isolated and purified from peripheral blood, stained with FITC- or PE-conjugated antihuman monoclonal antibodies (mAbs), in conjunction with anti-human FoxP3 for the enumeration of Treg cells, and quantified by flow cytometry. PE conjugated monoclonal CD34 (BI-3C5), FITC conjugated monoclonal CD45 (HI30), PE conjugated Monoclonal CD14 (TuK4), and FITC conjugated monoclonal CD16 (3G8) mAbs were used; all antibodies were purchased from Invitrogen (Grand Island, NY). Peripheral blood lymphocytes were stained with BioLegend’s True-Nuclear™ Human Treg Flow™ Kit (FOXP3 Alexa Fluor® 488/CD4 PE Cy5/CD25 PE) and were isolated by AutoMACS cell sorter (Miltenyi Biotec). All flow cytometry analysis was performed on a FACSCalibur (BD Biosciences) with BD CellQuest analysis software.
Walking performance was measured using the Timed 25 Foot Walk (T25FW). The protocol involved two trials of walking 25-feet as fast, but safely, as possible. We expressed T25FW performance as ft/sec based on an average of the two trials. This study included a measure of hand-grip strength, wherein participants were asked to squeeze a hand dynamometer (Jamar Hydraulic Hand Dynamometer, Lafayette Instruments, Lafayette, IN) as hard as possible. Participants were given 3 trials per hand with a 1-minute rest between trials, and the highest recorded value was taken as maximal grip strength (kg). Cognitive function was assessed using the Brief International Cognitive Assessment for Multiple Sclerosis (BICAMS) battery (Langdon DW, 2012) that includes the oral version of the Symbol Digit Modalities Test (SDMT), the California Verbal Learning Test-II (CVLT-II) and the revised Brief Visuospatial Memory Test (BVMT-R). The SDMT is a timed thinking task that involves associating numbers with symbols; the range of scores is 0-110. The CVLT-II is a test of verbal memory and involves recalling a list of 16 words read aloud by the examiner; the range of scores is 0-90. The BVMT-R is a test of visual memory that involves recalling and manually drawing a series of 6 abstract designs; the range of scores is 0-36. Mood was measured by the 30-item version of the Profile of Mood States (POMS) using a seven-day response set for capturing mood during the past week. We expressed the POMS score by summing the items into Total Mood Disturbance (TMD; 0-80) with lower scores reflected a better overall mood state.
All outcome assessments, data processing and analysis were conducted in a blinded fashion. Participants received remuneration for completing the measurements and for completion of the intervention sessions. Safety and adverse events were assessed at each visit by the study coordinator and were monitored monthly by the UIUC IRB committee.
Data were analyzed using SPSS version 24.0. Results are reported as mean and standard deviation (SD), unless otherwise noted (e.g., percentages, frequencies, medians). We compared initial differences between conditions using independent samples
We contacted 40 individuals with relapsing MS about the study, and 35 underwent screening for inclusion. Of these 35 individuals, 18 satisfied inclusion criteria, and 17 were enrolled and scheduled for baseline testing. There were 4 individuals who subsequently withdrew. The remaining 13 individuals underwent baseline testing and were randomized into an intervention condition (UVB phototherapy or placebo control). One person randomized into the phototherapy condition withdrew because of a family event. The final sample completing the study included 12 subjects with relapsing MS who were evenly distributed between treatment conditions (i.e., UVB Intervention, n=6; placebo-controlled condition, n=6); see the trial profile in
The descriptive characteristics of the study participants are provided in
Characteristic | UVB-Intervention Condition (n=6) | Placebo-Controlled Condition (n=6) | Test Statistic | ||
Age (yr) | 49.2 (14.8) | 55.0 (9.6) | .44 | ||
Height (cm) | 167.1 (6.5) | 164.8 (6.8) | .57 | ||
Weight (kg) | 70.0 (18.3) | 76.3 (17.7) | .56 | ||
Sex (n/% female) | 5/83% | 6/100% | .30 | ||
Skin type (n/%) | |||||
II | 1/17% | 0/0% | .34 | ||
III | 1/17% | 3/50% | |||
IV | 4/66% | 3/50% | |||
Assistive Device (n/% none) | 6/100% | 5/83% | .30 |
Classifies a person’s complexion and his or her tolerance of sunlight. II: white; burns
easily and tans with difficulty. III: cream white; sometimes mild burn and gradually tans. IV:
beige olive; rarely burn and tans easily.
The groups did not differ in skin type nor use of an assistive device for ambulation based on chi-square tests. Pre-study blood draws revealed that 25% of our study subjects were vitamin D deficient, 25% were vitamin D insufficient, and 50% had adequate vitamin D values. Vitamin D deficiency, insufficiency and adequate vitamin D was defined as 25(OH)D values less than 40 nmol/L, 25(OH)D values falling between 40-70 nmol/L and 25(OH)D values greater than 70nmol/L, respectively.At the completion of the study, 100% of the UVB intervention study subjects had adequate vitamin D levels as compared to only 50% of the placebo condition subjects. (see
Total Study Subjects | UVB Intervention Condition | Placebo Controlled Condition | |||||||
Vitamin D Level | Pre | Mid | Post | Pre | Mid | Post | Pre | Mid | Post |
Adequate (>70 nmol/L) | 50% | 58% | 67% | 33% | 67% | 100% | 67% | 50% | 50% |
Insufficient (40-70 nmol/L) | 25% | 25% | 17% | 50% | 33% | 0% | 0% | 17% | 17% |
Deficient (<40 nmol/L) | 25% | 17% | 17% | 17% | 0% | 0% | 33% | 33% | 33% |
The mixed-factor ANOVA identified a statistically significant condition by time interaction on serum vitamin D levels (
The descriptive and inferential statistics for the primary and secondary outcomes are provided in
Outcomes | Variable | UVB Intervention Condition (n=6) | Placebo-Controlled Condition (n=6) | ||||
Pre | Mid | Post | Pre | Mid | Post | ||
Primary | Vitamin D (nmol/L) | 71.7 (37.9) | 100.4 (51.6) | 124.7 (63.1) | 76.5 (41.1) | 62.5 (26.8) | 70.1 (34.7) |
Secondary | T25FW (ft/sec) | 5.2 (1.3) | 5.3 (1.3) | 5.5 (1.3) | 4.6 (2.5) | 5.4 (2.6) | 5.0 (2.6) |
Strength (kg) | 33.8 (14.7) | 33.5 (11.3) | 32.3 (9.8) | 29.2 (6.2) | 28.0 (8.6) | 25.8 (6.5) | |
SDMT (0-110) | 55.0 (22.1) | 59.0 (23.7) | 54.0 (13.9) | 54.5 (14.1) | 57.7 (13.5) | 57.7 (12.7) | |
CVLT (0-90) | 60.3 (7.6) | 55.2 (10.9) | 60.0 (4.7) | 54.3 (9.8) | 57.3 (8.8) | 58.3 (14.1) | |
BVMT-R (0-36) | 21.9 (7.4) | 25.7 (4.5) | 29.2 (4.7) | 19.2 (7.9) | 19.3 (6.5) | 23.8 (7.6) | |
TMD (0-80) | 36.5 (12.2) | 30.0 (15.1) | 32.0 (14.5) | 32.3 (6.9) | 25.0 (10.7) | 26.7 (12.7) |
Note. Values are mean score (standard deviation); T25FW = Timed 25-Foot Walk; SDMT = Symbol Digit Modalities Test;
CVLT = California Verbal Learning Test; BVMT-R = revised Brief Visuospatial Memory Test; TMD = Total Mood Disturbance.
Outcomes | Variable | Time | Condition | Time by Condition | |||
F-value | p-value | F-value | p-value | F-value | p-value | ||
Primary | Vitamin D | 4.32 | <.05 | 1.52 | .25 | 7.14 | <.005 |
Secondary | T25FW | 2.57 | .10 | 0.13 | .73 | 0.21 | .81 |
Strength | 2.55 | .10 | 0.98 | .35 | 0.35 | .71 | |
SDMT | 1.49 | .26 | .01 | .95 | 0.78 | .47 | |
CVLT | 0.73 | .50 | .14 | .71 | 1.39 | .27 | |
BVMT-R | 7.52 | <.005 | 2.05 | .18 | 0.72 | .50 | |
TMD | 2.39 | .12 | 0.65 | .44 | 0.02 | .98 |
UVB Intervention Condition (n=6) | Placebo-Controlled Condition (n=6) | |||
Variable | Pre | Post | Pre | Post |
Circulating progenitor cells(CD34+/CD45dim) | 0.17(0.13) | 0.17(0.12) | 0.11(0.05) | 0.28(0.34) |
Intermediate Inflammatory monocytes (CD14+/CD16+) | 1.07(0.63) | 1.58(0.68) | 1.83(1.66) | 1.83(1.73) |
Classical Phagocytic monocytes (CD14+/CD16-) | 2.98(1.41) | 5.66(3.46) | 4.02(3.04) | 4.05(4.28) |
Regulatory T cells -Tregs (CD4+/CD25+/FoxP3+) | 5.54(3.74) | 8.52(4.89) | 9.79(6.15) | 9.73(4.84) |
Note. Values are mean score (standard deviation). There are no significant interactions or main effects for the immune variables based on the 2-way ANOVAs.
Individual longitudinal changes in classical monocyte and Treg numerical status profiles appear different between the UVB treated subjects versus the placebo control group. For subjects 1, 3 and 6 a profile of increased intermediate monocyte, classical monocyte and Treg numerical status following UVB intervention occurred. The differences however, are not statistically significant, and the changes are not uniform across all UVB - treated subjects see
Although this study was unable to show significant UVB-induced MS immunoregulation or neurological improvement over the duration of the study, we do report some compelling observations. The UVB phototherapy condition, unlike the placebo-controlled condition, resulted in a statistically significant increase in serum 25(OH)D levels over the course of the 8-week trial. And while the changes in CPC, classical and intermediate monocyte and Treg cell counts following UVB intervention were not significantly different from those observed in the placebo control group, individual Treg cell profile changes following UVB light treatment, including a marked increase in Treg counts in 4/6 UVB treated individuals versus 1/6 of the placebo treated subjects was noted. This may reflect subject response variability and warrants further investigation. The trend towards an increase in Tregs in this population following UVB treatment represents an exploitable therapeutic potential given that Tregs play a critical role in the active suppression of autoimmunity. MS patients have been found to have lower circulating Treg cell numbers compared to healthy controls,
The observed variations in classical monocyte levels relative to alterations in intermediate monocyte levels may also indicate differences in disease duration, activity, or recovery. Classical monocytes are thought to primarily play a phagocytic role, whilst intermediate monocytes are thought to be associated with an inflammatory response. An increase in intermediate monocytes could be a differentiating factor between acute and chronic inflammation.
These results complement and extend those reported in a previous study of UVB persons with MS.
The limitations of the current pilot RCT include the small sample size, the short time course of the study, and the fact that the participants in our study were not all vitamin D deficient at baseline. Additionally, we did not consider disease duration or concurrent therapeutic interventions (medication) in our analysis. These factors did not preclude the demonstration of an intervention effect on serum vitamin D, but undermined our capacity to confidently interpret the UVB-induced changes in CPC, intermediate and classical monocyte, and Treg cell populations, and likely limited our ability to detect differences in secondary clinical end-points. Clinical benefits of UVB phototherapy may be more pronounced in patients with MS that have had a history of chronic vitamin D deficiency. This is important considering the evidence for vitamin D insufficiency and deficiency in MS,
Our observations have potential therapeutic implications, as the observed immunopathological heterogeneity in MS is likely to be both patient- and context-dependent, and may reflect either different host genetic factors that define different immune-mediated inflammatory responses, and the subsequent susceptibility to neuronal injury, or distinct triggers, respectively.
Given the report that the prevalence of MS in the US has been on a steady rise over the past 5 decades
The results presented in this study support the need for larger UVB phototherapy trials of longer duration to further investigate the Treg inductive capability of UVB radiation. In the future, we hope to evaluate UVB –induced immune profile changes, and their relative impact on MS disease course and relapse rate, in relation to vitamin D status. The individual longitudinal changes in the monocyte and Treg numerical status following UVB phototherapy shown here, highlight the necessity of individualized MS therapeutics, and the need for sophisticated computational strategies to aid in complex multivariable patient data analysis. Interventional studies designed to determine whether UVB phototherapy results in immunomodulation and subsequent amelioration of MS disease activity are justified.
We do not advocate chronic excessive sunlight exposure as a MS therapy, because of the associated increased risk of nonmelanoma skin cancer. We do however, support the recommendation for sensible sun exposure, as defined as 5-10 minutes of exposure of the arms, hands, legs and face 2 to 3 times per week, and increased dietary and supplementation of vitamin D intake to help ensure vitamin D sufficiency. This is particularly important in the context of the growing concern about vitamin D deficiency being an unrecognized epidemic among both children and adults in the US.
The results of this pilot trial indicate that UVB phototherapy represents a novel, safe, inexpensive and effective, nonpharmacological approach to increasing vitamin D levels in MS. UVB phototherapy corrects vitamin D deficiency. At the completion of the study, 100% of the UVB treated condition subjects had adequate vitamin D levels as compared to only 50% of the subjects in the placebo controlled condition (see
RM and CBP wrote the paper. BJR, BP, GN, MDL, SS and RM conducted the research. RM and CBP conducted the data analysis.
This study was funded by a grant from Sun Health Technologies. BJR, the studies neurologist is also the chief scientific officer for Sun Health Technologies. Thus, the University of Illinois created a conflict management plan for minimizing bias and undue influence in the running of the trial and data interpretation by triple blinding the study, and by restricting control of the study and access to the data to the University of Illinois primary investigator. The funding source aims to develop propriety phototherapy devices. The company did not play
RM and CBP wrote the paper. BJR, BP, GN, MDL, SS and RM conducted the research. RM and CBP conducted the data analysis.
Sun Health Technologies
This study was supported by a grant from Sun Health Technologies