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Marked improvement of thyroid function and autoimmunity by Aloe barbadensis miller juice in patients with subclinical hypothyroidism

Some natural compounds decrease serum levels of thyroid autoantibodies, but results are inconsistent and thyroid function has been evaluated infrequently; moreover, the effects of Aloe on thyroid autoimmunity and function have been examined in very few studies. This study stems from the observation of one co-author, who has Hashimoto’s thyroiditis (HT)-related subclinical hypothyroidism (SCH). Upon checking her biochemical thyroid panel when taking daily Aloe barbardensis Miller juice (ABMJ) for thyroid-unrelated reasons, she noticed a decrease in serum thyroperoxidase autoantibodies (TPOAb) and thyrotropin (TSH) and an increase in serum free thyroxine (FT4). Based on this observation, we enrolled 30 consecutive HT women with levothyroxine-untreated SCH and high TPOAb levels. All of them took ABMJ (50 ml daily) for nine months and were tested for serum TSH, FT4, free triiodothyronine (FT3) and TPOAb. Measurements were performed at baseline and at months 3 and 9. TSH, FT4 and TPOAb improved significantly already at month 3 and further (−61%, +23% and −56%) at month 9. However, FT3 decreased significantly at month 3 (−16%) with no further decrease at month 9, so that the FT4:FT3 ratio increased significantly (+33% and + 49%). At baseline, 100% of women had TSH > 4.0 mU/L and TPOAb > 400 U/ml, but frequencies fell to 0% and 37%, respectively, at month 9. In contrast, a control group (namely, 15 untreated SCH women of comparable age and baseline levels of TSH, FT4, FT3 and TPOAb) had no significant changes in any index. We conclude that the daily intake of 100 ml ABMJ for 9 months in women with HT-related SCH decreases the burden of thyroid autoimmune inflammation. In addition, ABMJ rescues thyrocyte function, with decreased need for conversion of the prohormone T4 into the more active T3 through ABMJ-induced inhibition of T4 deiodination.


Introduction
Aloe is a very old plant with medicinal properties that was discovered by the ancient Egyptians, who called it "the plant of immortality" [1]. The botanical name of Aloe vera is Aloe barbadensis Miller. It belongs to the Asphodelaceae (Liliaceae) family, and is a shrubby or arborescent, perennial, xerophytic, succulent, pea-green color plant. It grows mainly in the dry regions of Africa, Asia, Europe and America [1]. Records of its use were engraved in tablets thought to be from 2100 B.C. Aloe then travelled to various parts of the globe, and starting from the 17th century it had become a common medicinal plant [2]. The name Aloe vera derives from the Arabic word "Alloeh" meaning "shining bitter substance" and from the Latin word "vera" meaning amino acids, a dozen vitamins (A, B1, B2, B3, B5, B6, B12, C, E, choline, and folic acid), active enzymes (alkaline phosphatase, amylase, bradykinase, carboxypeptidase, catalase, cellulase, lipase, and peroxidase), anthraquinones (mostly known for their laxative effects), sterols, lignin, saponins, salicylic acids and others [1]. Interestingly, the thyroid hormone-forming tyrosine is the rarest amino acid present in Aloe Barbadensis leaves (28 µmol/100 g), while arginine is the main one (449 µmol/100 g) [3]. The anti-oxidant and/or anti-inflammatory properties of Aloe vera are explained by its content in vitamin A, C and E, in the glycoprotein C-glucosyl chromone, in certain sterols (campesterol, β-sitosterol and lupeol), vegetal hormones (auxins and giberrelins) and bradykinase. Campesterol, β-sitosterol, lupeol and two anthraquinones (aloin and emodin) act as analgesics and antiseptics [1]. These sterols are also found in shea butter and sabal/saw palmetto (Serenoa repens) [4,5]. Further details can be found in the Volume 1 of WHO (World Health Organization) Monographs on Selected Medicinal Plants [6]. As reviewed elsewhere [1], some uses of Aloe vera are based on scientific evidence in humans and/or animals, while other uses are based on tradition. Scientific-based therapies for Aloe vera include seborrheic dermatitis, psoriasis vulgaris, genital herpes, skin burns, wound healing, pressure ulcers, mucositis, radiation dermatitis, acne vulgaris, lichen planus, frostbite, aphthous stomatitis, type 2 diabetes mellitus, HIV infection, cancer prevention, ulcerative colitis and constipation. Traditional-based therapies for Aloe vera include alopecia, bacterial and fungal skin infections, parasitic infections, chronic leg wounds, systemic lupus erythematosus, arthritis and tic douloureux.
A number of natural compounds/nutraceuticals are being used to treat autoimmune thyroid diseases (AIT), namely Graves' disease (GD), GD-associated ophthalmopathy, HT and postpartum thyroiditis (PPT) . Though an exhaustive list of these substances goes beyond the scope of the present paper, examples of them include omega-3-fatty acid-rich small oily fish [7,8], L-carnitine [9][10][11], selenium [12][13][14][15][16][17][18][19][20][21][22][23][24][25][26][27][28][29], and myoinositol [27][28][29]. However, the observation that prompted the present study stemmed from a fortuitous observation (Table 1). One of the authors of the present paper, who has a history of HT-associated initial, mild hypothyroidism (also called subclinical hypothyroidism [SCH]), decided to take Aloe Barbadensis Miller juice (ABMJ), at the dose of 50 ml every morning on an empty stomach, as a skin soother and laxative. No other medications or over-the-counter compounds were taken. She checks thyroid function and thyroperoxidase autoantibodies [TPOAb] semiannually, since in one relative progression to overt hypothyroidism was preceded by a frank fall in FT4 and increase in both TSH and TPOAb approximately six months earlier. At the biochemical check performed three months after having started taking ABMJ, she was struck by the remarkable improvement of all indices ( Table 1). The improvement was even more impressive six months later (Table 1).
Based on this positive experience, we decided to test the effects of ABMJ administration in HT women with levothyroxine-untreated SCH and high levels of TPOAb.

Materials
The ABMJ taken by all patients was Aloe Vera 2 by ZUCCARI (Trento, Italy). Each 100 ml preparation of ABMJ, which has an energetic value of 7.40 kcal (31.20Kj), contains nonpasteurized and noncarbon filtered Aloe vera leaf juice and pulp (49.8 g), 0.2 g fats (0% saturated), 1.2 g carbohydrates, zero proteins, and 0.06 g minerals (of which, 0.02 g sodium). The juice also contains citric acid as an acidifier and sodium benzoate and potassium sorbate as typical preservatives with biocidal properties. Noteworthy, Aloe Vera 2 is free of aloin, a substance primarily contained in the outer cuticles of the leaves that is irritating to the intestinal mucosa. Once opened, the one-liter bottle has to be stored in the refrigerator, as recommended by the producer.

Patients
Based on the observations summarized in Table 1, we aimed to recruit women, aged 30 to 55 years and with HT-associated SCH (TSH > 4.0 mU/L; high levels of TPOAb), who had never been treated with L-T4 and/or supplements. In addition to past and current treatment with L-T4 and/or supplements, exclusion criteria were: (i) concurrent diseases, including diabetes mellitus and other autoimmune diseases; (ii) use of any nutraceuticals/drugs that affect the hypothalamic-pituitary-thyroid axis and autoimmunity. These criteria were verified with the family physicians.
Patients were informed of the aforementioned initial observation and enrolled upon signing an informed consent form. Thirty women, aged 20 to 55 years, were enrolled and all of them completed the study. They were treated with 50 ml ABMJ (Aloe Vera 2 ), which was taken in the morning on an empty stomach. To minimize confounding factors, all patients were directed to the same natural product health store where the said co-author bought ABMJ. The natural product health store personnel informed us that each patient had bought enough bottles to complete the study. Paralleling the initial observation, the duration of study was 9 months, with serum TSH, FT3, FT4 and TPOAb measured at baseline (time zero), and 90 ± 3 days (3 months) and 180 ± 3 days (9 months) later. Measurement of FT3 was added for sake of completeness.
For the purpose of comparison with a group of Hashimoto's thyroiditis women who were under no thyroid hormone replacement therapy and no supplementation with nutraceuticals, we took advantage of a database on patients with Hashimoto's thyroiditis (Interdepartmental Program of Molecular & Clinical Endocrinology and Women's Endocrine Health, University hospital of Messina). Based on age and levels of the fundamental indices (serum TSH and TPOAb) at time zero, the 30 Aloe-treated women could be matched to 15 women with SCH of comparable age (21-57 years) and baseline levels of serum TSH and TPOAb that had been measured with the same corresponding kits.

Statistics
Data are given as mean ± standard deviation (SD) and median. Differences between m ± SD of continuous variables were analyzed by ANOVA or by the Wilcoxon signed rank test for the indices with nongaussian distribution (TSH and TPOAb). Differences between proportions were analyzed by the two-tailed Fisher's exact test. The threshold for significance was set at a P value of < 0.05.

Biochemical thyroid data
Data are summarized in Tables 2 and 3, and illustrated in Figs. 1 and 2.
In the control group (Table 2), all biochemical indices were statistically similar to the corresponding ones of the study group. In addition, they did not change significantly over time.
In sharp contrast, in the study group all indices changed significantly at the first evaluation (end of the third month) ( Table 2). At this first post-treatment evaluation, serum TPOAb decreased by onefourth, while FT4 increased by one-sixth and FT3 decreased by the same magnitude. As expected from the well-known linear-log relationship in the thyroid hormone-driven feedback regulation of TSH secretion, serum TSH changed in the expected direction (decrease) and to a greater extent (3 times more) than the 15% increase in serum FT4. The 3-fold greater change held six months later, in response to the further increase in serum FT4 (approximately 60% compared to 24% increase in serum FT4); TPOAb levels continued to fall. Changes of serum TPOAb, TSH and FT4 augmented over time, whereas FT3 did not decrease further at month 9 compared with month 3. As a result, the FT4:FT3 ratio continued to increase (Table 2). Figs. 1 and 2 show that, both in the patients and in the control group, changes for each given index were uniform, with virtually no outliers.
In brief, all the changes observed are consistent with an amelioration of the intrathyroid autoimmunity process, with rescue of T4 synthesis by the thyrocyte (and subsequent decline in serum TSH) and simultaneous impairment of the deiodinase-mediated peripheral T3 formation. Table 3 shows that euthyroidism (TSH ≤ 4.0 mU/L) was achieved in 83% of patients already at the end of the third month of treatment. This proportion becomes exactly one-third (33%) using the very stringent criterion of TSH ≤ 2.5 mU/L. At the end of the ninth month, 100% or 83% of women were euthyroid, depending on TSH threshold. In contrast, all control women retained their SCH status, with a TSH > 4.0 mU/L (data not shown).
The bottom part of Table 3 shows that the worst biochemical scenario based on the combination of three indices (TSH, FT4 and TPOAb) occurred at baseline in 40 or 60% of patients (depending on the TPOAb threshold used). However, already by the third month of treatment the scenario improved at the best possible level (disappearance) or improved significantly.
None of the 30 women complained of any side-effects during the nine months of study.

Discussion
As checked with a PubMed search on September 30, 2017 using the string "Aloe and thyroid", except for a letter to the Editor reporting on a single patient [30] and one paper on male mice [31], there are no other studies describing the effects of Aloe on thyroid function. A 56-yr-old woman with lichen planus, who refused treatment with corticosteroids, took Aloe vera juice (10 ml/day) for 11 months [30]. No improvement Table 2 Changes observed in the specified biochemical thyroid indices (all analyzed as continuous variables) during and at the end of a 9-month duration administration of Aloe vera barbadensis Miller juice in 30  of lichen occurred. Serum FT3 was barely low, FT4 at the lower reference limit and TSH normal. After discontinuation of Aloe, thyroid hormone levels were reported to be normal. Taking into account the few data provided and not having mentioned the presence of lymphocytes in the thyroid smear, the hormone picture is consistent with central hypothyroidism or with the nonthyroid illness syndrome, rather than primary hypothyroidism. In the mice study by Kar et al. [31], groups of seven male mice were treated, for 15 days, with extracts of Aloe vera (125 mg/kg) or other plants by gastric intubation. Compared with administration of vehicle (control), a significant increase in serum T4 concentration was observed in mice treated with Aegle marmelos or Bacopa monnieri (+31% or +41%, P < 0.01 for both), while T3 concentration decreased in the former group only (−62% [P < 0.001] or +5.7%). In Aloe veratreated animals, both T4 and T3 levels decreased significantly (−25% [P < 0.01] and −13% [P < 0.05]). No mechanistic explanation was provided [31].
Here we have shown that the fortuitous observation of decreased serum TSH and TPOAb in association with an increased concentration of serum FT4 after a 9-month administration of Aloe vera juice was confirmed in a cohort of women with the same thyroid disorder: HTrelated SCH. We also measured serum FT3 and found a decline after 3 months only, with no further decline after another six months. In sharp contrast, untreated SCH women of comparable age and baseline levels of TSH, FT4, FT3 and TPOAb had no significant changes in any index.
Similarly to Nordio and Basciani [28], we complemented measurement of serum FT4 with that of serum FT3 in our subclinically hypothyroid HT patients. Unlike the 4.5% increase in serum FT3 following a 6-month supplementation with selenomethionine plus myoinositol [28], we found a 16% decrease after 3 months of supplementation with Aloe vera, and this value did not change in either direction after another 6 months of supplementation (Table 2). Consequently, the FT4:FT3 ratio (pmol/L:pmol/L) increased from 1.83 at baseline to 2.47 (+35%) at month 3 and 2.78 (+52%) at month 9 after Aloe vera supplementation, as did, but to a lower extent (2.95 to 3.21 [+9%]), after 6 months of supplementation with selenomethionine plus myo-inositol [28]. Upon studying in vitro other HT patients' circulating lymphocytes that were cultured in the presence of selenomethionine plus myo-inositol, our group has found a dose-dependent decline in the extracellular concentration of the hydrogen peroxidestimulated lymphocyte-secreted chemokines (CCL2, CXCL9 and CXCL10) [32]. Because (i) circulating lymphocytes can be a proxy of the lymphocytes that infiltrate the thyroid in HT, (ii) the said chemokines are markers of functional damage of the thyrocyte [33,34], and (iii) increased peripheral deiodination of T4 into the more potent T3 is a compensatory mechanism for the diminished secretion of T4 from the thyroid gland, we propose the following interpretation for the amelioration of the thyroid autoantibody profile and hormone profile induced by either selenemethionine plus myo-inositol or Aloe vera juice. Upon supplementation, the decreased intrathyroid inflammation and decreased chemokine-mediated damage of the thyrocyte result in greater T4 secretion and decreased need to generate T3 in the periphery compared to the pre-supplementation period.
The benefit provided by the Aloe vera juice can be measured on a clinically relevant scale, namely return to the euthyroid status. Before supplementation, 100% of our patients were subclinically hypothyroid (TSH > 4.0 mU/L), but nine months after supplementation none was so. If the TSH threshold is lowered to 2.5 mU/L (which is relevant in the setting of the first trimester of gestation), after 3 and 9 months of Aloe vera supplementation two-thirds and one-sixth of women would have the undesirable level of TSH > 2.5 mU/L. Remaining in the obstetrical setting, the decline in TPOAb from the first week of gestation through 12 months postpartum upon supplementing Italian women with selemethionine for all this time was beneficial [26], in that the decline of TPOAb (a well-known risk factor for PPT development) was followed by a 2-fold lower rate of PPT and hypothyroidism compared to the   untreated group. We speculate that the same outcome would very likely occur should women take Aloe vera juice for the entire duration of gestation and the first 12 months after delivery.
In conclusion, a 9-month treatment with 50 ml/day of Aloe vera juice restores euthyroidism in 100% of HT-related initial/mild hypothyroidism (also known as subclinical hypothyroidism), with appreciable results already evident after 3 months, and markedly decreases their highly elevated serum TPOAb levels (again already evident after 3 months). These beneficial results are comparable to, if not greater than, those reported in the literature for selenium alone or selenium associated with myo-inositol. In addition to an ameliorative effect on thyroid inflammation and subsequent rescue of the hormone biosynthesis by the thyrocytes, Aloe vera juice is likely to act on the peripheral deiodination of T4 into T3. Randomized, double-blind placebocontrolled trials are warranted.

Declarations of interest
None. This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.