Coronavirus disease 2019 (COVID-19) has been associated with manifestations in the post-viral stage that affect multiple organ systems (1). Emerging literature has reported residual effects of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection manifested by a wide spectrum of symptoms that persist or begin after the acute-viral stage called “long-term” of COVID-19 (2–4), “long COVID” (5, 6), or “post-COVID-19 syndrome” (7, 8). According to the guideline (9), “acute COVID-19” represents signs and symptoms for up to 4 weeks, while “post-COVID-19 syndrome, long-term or long COVID” comprises signs and symptoms that develop during or after an infection consistent with COVID-19 present for more than 12 weeks and is not attributable to alternative diagnoses (4, 9).

Persistent symptoms in survivors of COVID-19 who had been hospitalized tend to vary according to the time of assessment. Carfi et al. (10) showed a high proportion of individuals who still reported fatigue (53.1%), dyspnea (43.4%), joint pain (27.3%), and chest pain (21.7%) 2 months after COVID-19 symptom onset. In a prospective cohort of patients interviewed by telephone after 4 months of hospital discharge by COVID-19, 51% reported the presence of at least one symptom that did not exist before the disease, being fatigue (31%), cognitive symptoms (21%), and new-onset dyspnea (16%) the most frequent manifestations (3).

Survivors of COVID-19 who had been hospitalized still present long-term health consequences at 6 months (2) or 1 year (11) after recovering from the acute phase of SARS-CoV-2 infection. Fatigue or muscle weakness (63%), sleep difficulties (26%), and anxiety or depression (23%) were the most common clinical manifestations after 6 months of symptom onset (2). A case report of a 50-year-old female nurse described a history of fatigue on minor exertion and persistent dysphonia without organic alterations after 1 year of mild COVID-19 (11).

Among numerous efforts to discover adjuvant therapeutic compounds against SARS-CoV-2 infection (12–14), it has been postulated through the effective increase in serum 25-hydroxyvitamin D (25[OH]D) levels after a single, safe high dose of vitamin D₃ used in our study (15) the possible attenuate of the persistence or onset of symptoms. According to a previous systematic review (16), a single high dose of vitamin D₃ significantly reduced levels of inflammatory cytokines (interleukin-6 and tumor necrosis factor-alpha, TNF-α) (17), improved endothelial function in patients with a history of stroke (18), type 2 diabetes (19), and cardiovascular function (20). Additionally, a single high dose of vitamin D3 promoted a good functional modified ranking scale with decreased mortality risk in ischemic stroke survivors with baseline serum 25(OH)D < 30 ng/ml at 6 months (21), while high concentrations of serum 25(OH)D were longitudinally associated with better quality of life (QoL) and less fatigue within 2 years after treatment (22).

Serum 25(OH)D levels are expected to remain sufficient for up to 4 months (23) following a single high dose of vitamin D₃ of approximately 200,000 IU, even at baseline 25(OH)D levels 20 ≤ ng/ml (16). The main hypothesis was that a single high dose of vitamin D₃ would sustain long-term serum 25(OH)D levels aiding in the modulation of inflammation, and innate and adaptative immune responses related to post-COVID-19 manifestation. In view of the biological plausibility and presumed benefit of vitamin D mitigating the persistent symptoms of COVID-19, this study investigates the reported persistent or new symptoms 1 year after a single high dose of vitamin D₃ and hospitalization in patients with moderate to severe COVID-19.

Of the 1,240 patients assessed for eligibility, 240 underwent randomization during the acute phase of SARS-CoV-2 infection, with 120 assigned to each group. Of the 110 patients discharged in the vitamin D₃ group, 21 were excluded due to the absence of a telephone number, 8 were excluded due to lack of contact, 8 withdrew consent, and 2 did not receive vitamin D₃. Of the 112 patients discharged in the placebo group, 25 were excluded due to the absence of telephone numbers, 10 were excluded due to lack of contact, 3 died after hospital discharge, and 1 withdrew consent (Supplementary Figure 1). Overall, mean (SD) age was 54.3 (13.1) years, BMI was 32.4 (6.5) kg/m², 77 (53.5%) were men, 82 (56.9%) patients were White, 43 (29.9%) were Pardo, 18 (12.5%) were Black, and 1 (0.7%) was Asian (Table 1).

Fever demonstrated a significant main effect of time with a reduction from baseline to 6 months (52 to 0, P < 0.001) and from baseline to 12 months (52 to 0, P < 0.001) in the vitamin D₃ group compared to the placebo group from baseline to 6 months (53 to 0, P < 0.001) and from baseline to 12 months (53 to 0, P < 0.001) (Table 2). No significant difference between the vitamin D₃ and placebo groups for fever was observed at baseline (52 vs. 53, P = 1.00) (Table 2).

No significant differences between the vitamin D₃ and placebo groups for cough, fatigue, fever, myalgia, joint pain, runny nose, nasal congestion, sore throat, hypertension, diabetes, cardiovascular disease, rheumatic disease, asthma, chronic obstructive pulmonary, and chronic kidney disease were observed (Table 2). In addition, no significant differences between groups were found for QoL and frequency of new symptoms (Figures 1A,B), nor Kaplan-Meier curves for time manifesting symptoms (Figure 2A) and frequency of participants manifesting at least one symptom (Figure 2B) related to COVID-19 up to 1 year of follow-up.

In this post-hoc exploratory analysis from a multicenter, double-blind, placebo-controlled, randomized clinical trial, a single high dose of vitamin D₃ did not significantly differ from a placebo for COVID-19 symptoms, coexisting diseases, or QoL. To the best of our knowledge, this is the first randomized clinical trial to investigate the reported persistent or new symptoms in patients with moderate to severe COVID-19 1 year after a single high dose of vitamin D₃ and hospitalization.

Cough, fatigue, fever, myalgia, joint pain, runny nose, diarrhea, nasal congestion, and sore throat presented a significant main effect of time with reduced frequency from baseline to 6 and 12 months in both groups, except for the increase in chronic kidney disease, not differing between vitamin D₃ and placebo.

The findings demonstrated manifestation of at least one symptom related to COVID-19 persisting for up to 1 year after the acute phase of SARS-CoV-2 infection in 44.3% of the participants in the vitamin D₃ group and 48.9% in the placebo group. Furthermore, our results showed that from the first trimester after hospital discharge, at least 40% of participants did manifest any symptoms related to COVID-19 for both groups, in line with previous evidence (2, 3, 10), although we did not observe a significant difference in time to manifesting symptoms between the vitamin D₃ and placebo groups.

The most frequent symptoms at 6 and 12 months were joint pain (49.3 and 32.4%, respectively), fatigue (32.4 and 42.3%), and myalgia (21.1 and 38.0%) in the vitamin D₃ group compared to joint pain (49.3 and 27.4%), fatigue (38.4 and 38.4%), and myalgia (21.9 and 23.3%) in the placebo group. These findings are very similar to 53.1% fatigue and 27.3% joint pain at 2 months demonstrated by Carfi et al. (10), 31% fatigue at 4 months by the Comebac study (3), and 63% fatigue or muscle weakness at 6 months by Huang et al. (2).

Regarding new symptoms after hospital discharge for COVID-19, muscle weakness and memory impairment were among the most frequent in the vitamin D₃ group, whereas in the placebo group, in addition to the two aforementioned symptoms, a trend (P = 0.06) of greater hair loss was observed in comparison to the vitamin D₃ group, being the third most frequent manifestation.

The technical term for this hair loss is reported telogen effluvium. Telogen effluvium is characterized by diffuse hair loss within months of a significant systemic stressor, such as the premature follicular transition from the anagen (active growth phase) to the telogen (resting phase), the latter phase lasting approximately 3 months, after which excessive hair loss ensues (27). In fact, some evidence has pointed to SARS-CoV-2 infection as a precursor to acute telogen effluvium (28–32).

A research letter presented 10 patients with telogen effluvium following SARS-CoV-2 infection (28). All patients were women with a median age of 55, had laboratory-confirmed COVID-19, and had no history of hair loss. Mild symptoms were reported in six of them and severe disease in four, requiring hospitalization for an average of 7 days, and they all experienced excessive hair loss within months after infection, which included hair coming out in large clumps and thinning along the frontal hairline (28). These data corroborate other reports showing, on average, the onset of hair loss 50 days after the first symptom of COVID-19 infection (29, 31), associated or not with trichodynia (30), and may present duration ranging from 12 to 100 days (32).

One of the attributions to vitamin D functions would be related to hair loss. Vitamin D plays an important dermatological and dermatotherapeutic role in affecting the hair cycle due to its anti-inflammatory and immunomodulatory properties, and regulation of keratinocyte differentiation and proliferation (33). Recently, the benefit of oral vitamin D₃ (200,000 IU) therapy in patients suffering from diffuse hair loss (telogen effluvium) has been demonstrated (34). However, conclusive studies regarding the presumed benefit of vitamin D in hair loss are lacking. There are no clinical trials that have investigated the efficacy of vitamin D in managing hair loss.

With respect to the QoL, Jacobs et al. (35) reported persistent symptoms at 35 days after hospital discharge for COVID-19 infection associated with lower odds of rating QoL and its categories (general health, physical health, mental health, and social functioning). Similarly, Chen et al. (25) demonstrated a poor health-related QoL among COVID-19 patients, and those women were negatively associated with physical function, bodily pain, and emotional domain at the 1-month follow-up. Even 4 months after hospitalization for COVID-19, the median SF-36 was 46.9 (IQR, 31.2–68.8) for vitality and 57.5 (IQR, 40.0–75.0) for general health (3). Our results show a slight, but not significant, improvement at 6 months, although we have failed to observe differences between the vitamin D₃ and placebo groups.

During follow-up interviews, participants were also asked whether they had been using medications and/or supplements. Only 3 participants in the vitamin D₃ group and 2 in the placebo group reported using a commercially available multivitamin or vitamin D (∼200 IU). Statistical sensitivity analysis excluding the aforementioned participants showed no significant difference in persistence or new symptoms, and QoL reported up to 1 year of follow-up between the vitamin D₃ (n = 68) and placebo (n = 71) groups.

The postulated role of vitamin D in SARS-CoV-2 infection suggests action on the innate and adaptive immune system while mitigating excessive signaling for local and systemic inflammation (36). Our group failed to observe an effect of a single high dose of 200,000 IU of vitamin D₃ eliciting significant changes in systemic inflammatory cytokines, chemokines, and growth factors compared with placebo (37). Furthermore, this supplementation strategy was not able to significantly reduce the length of hospital stay compared to those who were supplemented with a placebo (15). Nevertheless, clinicians and researchers continue the search for supplements with therapeutic properties that effectively mitigate post-COVID-19 syndrome (38).

Aside from experimental design, the strength of this study includes being a prospective trial investigating the effects of vitamin D₃ on persisted or new symptoms after hospitalization for COVID-19, and enrollment of moderate to severe patients followed up for 1 year.

This study has several limitations. First, this investigation did not assess 25(OH)D concentrations at 6 and 12 months. Second, it was not feasible to evaluate the patients in person, and it was necessary to rely on the patients’ memory to obtain the data retrospectively. Third, regarding the new symptom onset related to COVID-19, it is not possible to determine when it started or ended. Fourth, at the time, most patients were admitted to the hospital’s emergency room, some of whom did not remember their telephone numbers, and the hospital’s social care staff did not locate the patients’ or family’s telephone numbers at hospital/study admission or during hospitalization; therefore, the authors are not able to state whether approximately 20% of participants excluded due to absence of telephone numbers would be alive after hospital discharge.

This study does not rule out the possibility that intermediate doses of vitamin D₃ administered during follow-up could imply different results or that a single high dose of vitamin D₃ followed by treatment with a recommended standard intake of at least 400 IU/day (39) may result in positive effects in favor of vitamin D and therefore could be evaluated in the future. Therefore, further studies are needed to prospectively assess the efficacy of fractionated high doses of vitamin D₃ on persistent or new symptoms after hospitalization for COVID-19.

Despite the trend toward greater hair loss (telogen effluvium) as the third most frequent new symptom in the placebo group, we did not rule out the possibility that vitamin D could prevent marked telogen effluvium if applied as a booster dose around 3 months after hospital discharge. Our findings advance in showing that a single high dose of vitamin D₃ did not support the initial hypothesis of an eventual role in innate and adaptive immune responses modulating long-term COVID-19, even in view of the expected significant reduction in COVID-19 symptoms from baseline up to 6 months and 1 year. Therefore, a single high dose of 200,000 IU of vitamin D₃ compared to placebo did not interfere with the persistence or new symptoms, and QoL reported by moderate to severe patients after hospitalization for COVID-19 up to 6 months and 1 year of follow-up.

The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation. Deidentified participant data of this study must be requested from the corresponding author RP, rosamariarp@yahoo.com. The codebook of this study will be made available upon request by qualified clinical researchers for specific purposes dependent on the nature of the request and the intention use of the data, with investigator support. The request must include a statistician. The lead author (RP) affirms that the manuscript is an honest, accurate, and transparent account of the study being reported; that no important aspects of the study have been omitted; and that any discrepancies from the study as originally planned (and, if relevant, registered) have been explained.

The studies involving human participants were reviewed and approved by the Ethics Committee of the Clinical Hospital of the School of Medicine of the University of São Paulo (approval numbers: 30959620.4.0000.0068). The patients/participants provided their written informed consent to participate in this study.

RP: full access to all the data in the study and responsibility for the integrity of the data and the accuracy of the data analysis and funding and supervision. AF, IM, and RP: concept and design. AF and RP: drafting of the manuscript and statistical analysis. LS, MS, and VC: administrative, technical, or material support. All authors: acquisition, analysis, and interpretation, and critical revision of the manuscript for important intellectual content.