Introduction: Metformin was found to reduce elevated levels of anterior pituitary hormones. Its thyrotropin-lowering effect was more pronounced in individuals receiving myo-inositol. The aim of the present study was to investigate whether the concomitant supplementation of myo-inositol determines the impact of metformin on prolactin levels. Methods: The study population consisted of two groups of women with mild-to-moderate hyperprolactinemia. Group 1 included 24 individuals receiving myo-inositol preparations (2 g daily for at least 6 months), while 24 inositol-naïve women belonged to group 2. Both groups were matched for age, insulin sensitivity, and prolactin concentration. For the following 6 months, all women were treated with metformin (1.7 daily). Plasma glucose levels, the homeostatic model assessment of insulin resistance ratio (HOMA-IR), glycated hemoglobin, as well as plasma levels of total prolactin, monomeric prolactin, thyrotropin, free thyroid hormones, adrenocorticotropic hormone, and insulin-like growth factor-1 were measured at baseline and after 6 months of metformin treatment. Results: Metformin reduced plasma glucose, HOMA-IR, and glycated hemoglobin in both study groups, but this effect was more pronounced in group 1 than group 2. Treatment-induced changes in total and monomeric prolactin levels were significant only in group 1. There were no differences between follow-up and baseline values of thyrotropin, free thyroxine, free tri-iodothyronine, adrenocorticotropic hormone, and insulin-like growth factor-1. Treatment-induced changes in prolactin concentration correlated with baseline prolactin levels, baseline values of HOMA-IR, and the impact of treatment on HOMA-IR. Discussion: The obtained results suggest that myo-inositol supplementation potentiates the inhibitory effect of metformin on prolactin levels in women with hyperprolactinemia.

© 2023 The Author(s). Published by S. Karger AG, Basel

Introduction

Recent studies have shown that metformin decreased circulating levels of anterior pituitary hormones: thyrotropin, gonadotropins (FSH and LH), and prolactin if baseline levels of these hormones were elevated [1-7]. The reduction in prolactin levels was observed in subjects with prolactin-secreting tumors [5], traumatic brain injury [5], drug-induced prolactin excess [6, 7], and empty sella syndrome [5]. No effect was found only if prolactin excess resulted from the presence of the high-molecular form of prolactin (macroprolactin) [8]. The prolactin-lowering effect of metformin correlated with the degree of hyperprolactinemia. This effect was stronger in postmenopausal women on hormone replacement therapy and young women taking oral contraceptive pills than in postmenopausal and premenopausal women not taking estrogens [9, 10]. In addition to monotherapy, metformin potentiated a prolactin-lowering effect of small doses of bromocriptine [11]. However, its addition to ongoing high-dose cabergoline treatment in patients with cabergoline-resistant prolactinomas failed to show a consistent inhibitory effect on circulating prolactin levels, and only 2 out of 10 patients were partial responders [12].

Myo-inositol is the most abundant form of inositol, and its derivatives play an important role in the signaling of various hormones, including insulin, thyrotropin, and gonadotropins [13]. Myo-inositol was also found to potentiate the effect of selenomethionine on plasma levels of thyrotropin and free thyroid hormones in patients with subclinical hypothyroidism and autoimmune thyroiditis [14, 15], as well as to enhance the impact of metformin on thyrotropin levels in women with mild thyroid hypofunction [16]. Moreover, administered together with folic acid, myo-inositol reduced plasma levels of LH, prolactin, testosterone, and the LH/FSH ratio [17]. Its prolactin-lowering effect was observed despite the fact that baseline concentrations of this hormone were within the reference range [17].

However, the interaction between metformin and inositol at the level of the pituitary gland is still very poorly understood. Therefore, the aim of the present study was to investigate whether concomitant use of myo-inositol determines the impact of metformin on prolactin levels.

Methods

After study approval by the Institutional Review Board, all participants provided signed informed consent to participate in the study. The study protocol followed the principles of the Declaration of Helsinki.

Patients

Young women (aged 18–45 years old) were eligible to participate in the study if they had mildly or moderately elevated plasma total prolactin levels (between 30 and 60 ng/mL [640 and 1,280 mIU/L]) found on two different occasions and if they met the criteria of prediabetes (fasting plasma glucose at least 100 mg/dL but less than 126 mg/dL and/or plasma glucose 2 h after a glucose load at least 140 mg/dL but below 200 mg/dL), despite complying with lifestyle modifications for at least 3 months preceding the study.

The exclusion criteria included prolactinoma, other pituitary tumors, macroprolactinemia (defined as the prolactin recovery below 40%), diabetes mellitus, thyroid, parathyroid or adrenal disorders, impaired renal or hepatic function, anemia, malabsorption syndrome, any other serious disorders, pregnancy or lactation, poor patient compliance, and concomitant use of dopaminergic agents or drugs known to interact with metformin or inositol.

The study population consisted of 2 groups of patients. Group 1 included 24 individuals receiving myo-inositol (2 g daily) for at least 6 months. The remaining 24 women (belonging to group 2) were selected from a group of 56 women meeting all inclusion and exclusion criteria but never receiving inositol preparations. The aim of this selection was to create two groups matched for age, insulin sensitivity, and prolactin concentration. A preliminary sample size analysis showed that the study population was enough and exceeded the required number of patients (21 per each group). Similar numbers of patients were recruited between June and August (n = 25) and between December and February (n = 23).

Study Design

All patients were started on metformin at an initial dose of 850 mg once daily. After 2 weeks, the dose was increased to the full dose of 850 mg twice daily. Moreover, group 1 continued myo-inositol treatment with the same dose as before (2.0 g daily). All participants were instructed to maintain their lifestyle habits during the course of the study. Patients were evaluated every 8 weeks until the end of study. Treatment adherence was assessed by the pill counting method. Adverse events were recorded throughout the study by direct questioning.

Laboratory Assays

Samples of venous blood were collected in the morning, after an overnight fast at study entry and 6 months later. In group 1, glucose homeostasis markers and total prolactin were also assessed before implementation of myo-inositol treatment. Samples were collected from the antecubital vein between 7:00 and 8:30 a.m. after an overnight 12h fasting and assessed in duplicate. Before blood collection, the participants had been resting in a temperature-controlled (24–25°C) quiet room for at least 30 min in the seated position. In order to perform polyethylene glycol precipitation, equal volumes (250 μL) of plasma and 25% cold polyethylene glycol 6000 dissolved in phosphate-buffered saline (Sigma, 137 mmol/L sodium chloride, 10 mmol/L sodium phosphate, pH = 7.4) were mixed and incubated for 10 min. After incubating for 10 min at room temperature and vortex mixing for 30 s, the suspension was clarified by centrifugation at 3,000 g for 30 min. To correct for the dilution with polyethylene glycol, the post-polyethylene glycol prolactin concentration was determined by multiplying the prolactin result by two. Glucose and glycated hemoglobin were assessed using the COBAS Integra 400 Plus analyzer (Roche Diagnostics, Basel, Switzerland). Plasma levels of prolactin before polyethylene glycol precipitation (total prolactin), prolactin in the supernatant after polyethylene glycol precipitation (monomeric prolactin), thyrotropin, free thyroxine, free tri-iodothyronine, and insulin were measured using acridinium ester technology (ADVIA Centaur XP Immunoassay System, Siemens Healthcare Diagnostics, Munich, Germany). Circulating levels of adrenocorticotropic hormone (ACTH) and insulin-like growth factor-1 were measured by solid-phase enzyme-labeled chemiluminescent immunometric assays (Immulite, Siemens, Munich, Germany). The homeostasis model assessment of insulin resistance index (HOMA-IR) was calculated by dividing the product of fasting glucose (mg/dL) and insulin (mIU/L) by a constant (405).

Statistical Analysis

Before statistical testing, all variables were logarithmically transformed to achieve a normal distribution. Between- and within-group comparisons were carried out by two-sample t tests and Student’s paired t tests, respectively. Nominal data were analyzed using χ2 test. Correlations were determined using Pearson’s r tests for two continuous variables; phi coefficient for one continuous and one categorical variable; and point-biserial for two categorical variables. The results were regarded as statistically significant if two-tailed p values adjusted for multiple testing were below 0.05.

Results

At study entry, there were no statistically significant differences between both groups in age, smoking, conditions leading to hyperprolactinemia, the body mass index (BMI), blood pressure, glucose, HOMA-IR, glycated hemoglobin, and circulating levels of total prolactin, monomeric prolactin, thyrotropin, free thyroxine, free tri-iodothyronine, ACTH, and insulin-like growth factor-1 (Tables 1, 2). In group 1, glucose, HOMA1-IR and glycated hemoglobin (but not BMI and prolactin) were lower at baseline than before implementation of myo-inositol treatment (online suppl. Table 1; for all online suppl. material, see www.karger.com/doi/10.1159/000528542). All patients complied with treatment and dietary recommendations. Metformin and myo-inositol were well tolerated. No significant adverse effects were reported, and there were no dropouts during the follow-up period.

Table 1.

Baseline characteristics of participants

Table 2.

Effect of metformin on BMI, glucose homeostasis markers, and hormone levels in myo-inositol-treated and myo-inositol-naïve women with hyperprolactinemia and prediabetes

In group 1, but not group 2, metformin tended to reduce BMI. Although metformin decreased plasma glucose, HOMA-IR, and glycated hemoglobin in both groups, this effect was more pronounced in group 1 than group 2. Only in group 1, the drug significantly reduced total and monomeric prolactin levels, while in group 2, the effect on prolactin levels did not reach the level of significance. There were no differences between follow-up and baseline values of thyrotropin, free thyroxine, free tri-iodothyronine, ACTH, and insulin-like growth factor-1. Both groups differed in follow-up values of glucose, HOMA-IR, glycated hemoglobin, and prolactin (Table 2).

At entry, there were correlations between prolactin and HOMA-IR (total prolactin: r = 0.41, p = 0.002; monomeric prolactin: r = 0.46, p = 0.001), between prolactin and thyrotropin (total prolactin: r = 0.29, p = 0.048; monomeric prolactin: r = 0.34; p = 0.026), as well as between prolactin and BMI (total prolactin: r = 0.35, p = 0.016; monomeric prolactin: r = 0.38, p = 0.008). The impact of treatment on prolactin levels correlated with baseline concentrations of this hormone (total prolactin: group 1: r = 0.47, p = 0.002, group 2: r = 0.42, p = 0.004; monomeric prolactin: group 1: r = 0.52, p < 0.001, group 2: r = 0.46, p < 0.001), baseline values of HOMA-IR (total prolactin: group 1: r = 0.35, p = 0.021, group 2: r = 0.31, p = 0.040; monomeric prolactin: group 1: r = 0.39, p = 0.004, group 2: r = 0.34, p = 0.028), and with treatment-induced changes in HOMA-IR (total prolactin: group 1: r = 0.40, p = 0.008, group 2: r = 0.37, p = 0.018; monomeric prolactin: group 1: r = 0.42, p = 0.003, group 2: r = 0.35, p = 0.018). In group 1, there were no correlations between the strength of metformin action on total and monomeric prolactin and on glucose homeostasis markers and total prolactin levels before myo-inositol supplementation, as well as between the strength of metformin action and the duration of myo-inositol treatment.

Discussion

Metformin administered alone only tended to reduce prolactin levels. There are two possible explanations for this finding. First, this may result from the fact that the participants had either mildly or moderately elevated concentrations of this hormone. For ethical reasons, individuals with marked hyperprolactinemia secondary to pituitary tumors were excluded because they always require treatment with dopamine agonists or surgery. In line with this explanation, the impact of metformin on prolactin levels correlated with baseline concentration of this hormone. According to the alternative explanation, the impact of metformin on pituitary hormone secretion is dose dependent. In previous studies, this effect was more pronounced if metformin was administered at the dose of 2.55–3 g daily than at lower doses [4, 6]. Despite higher metformin content in the pituitary than in other brain regions (the hippocampus, cerebellum, hypothalamus, olfactory bulbs, striatum, and frontal cortex) [18], relatively high doses of this drug needed for exerting its pituitary effects suggest the low sensitivity of pituitary cells to metformin.

The most important finding of the present study was that metformin used as an add-on treatment was superior to metformin alone in reducing prolactin levels. The dose of myo-inositol used in the current study (2 g daily) was the same as in the study by Artini et al. [17] but higher than that used by another research group (600 mg daily) [14, 15]. Unfortunately, there is no recommended daily allowance for inositol, and there is no standardized dosing schedule. The results of our study suggest that this beneficial effect of metformin and myo-inositol reflects pharmacodynamic interactions between the two agents. They cannot be explained by population heterogeneity because of strict inclusion and exclusion criteria, as well as because of matching both groups for age, insulin sensitivity, and prolactin levels. Moreover, the study groups did not differ in the percentage of women with drug-induced hyperprolactinemia, prolactin excess secondary to empty sella syndrome, and hyperprolactinemia resulting from traumatic brain injury. An eventual impact of greater insulin resistance before myo-inositol administration or of a time-dependent effect of myo-inositol does not also seem convincing because the strength of metformin action did not correlate with values of pre-myo-inositol HOMA-IR and with the duration of myo-inositol treatment.

Pituitary effects of metformin/myo-inositol combination therapy in the current study were limited to a decrease in total and monomeric prolactin levels, and the magnitude of this reduction correlated with baseline prolactin levels. However, there were no differences between baseline and follow-up concentrations of the remaining pituitary and downstream hormones: thyrotropin, free thyroxine, free tri-iodothyronine, ACTH, and insulin-like growth factor-1, all of which were within the reference range. These findings suggest that the impact of metformin/myo-inositol combination therapy is restricted to overactive pituitary cells, and the strength of this effect depends on the degree of their overactivity. In turn, the impact on function of intact anterior pituitary cells seems to be negligible, and therefore the combination therapy does not seem to pose a risk of inducing drug-induced hypopituitarism.

Between-group differences in the impact on prolactin levels were accompanied by different metabolic effects exerted by metformin alone or in combination with myo-inositol. Moreover, in both study groups, the decrease in prolactin levels correlated with baseline values of HOMA-IR, as well as with treatment-induced changes in HOMA-IR. These findings suggest that pituitary and metabolic effects of metformin and metformin/myo-inositol combination therapy are reciprocally related. To the best of our knowledge, the present study is the first one which compared the impact of metformin/myo-inositol combination therapy and metformin monotherapy on glucose homeostasis markers. However, head-to-head comparisons showed that in women with polycystic ovary syndrome, there were no differences in the impact on glucose homeostasis markers between metformin and myo-inositol [19] or that inositol was superior to metformin [20].

Hyperprolactinemia leads to weight gain and obesity/overweight through a reduction in insulin sensitivity, orexigenic actions on the central nervous system, promotion of positive energy balance, and control of adipocyte differentiation and fate, while the decrease in prolactin levels causes a reduction in body weight [21]. In the present study, metformin/myo-inositol combination therapy, but not metformin alone, tended to reduce BMI. This observation is in line with the results of a recent meta-analysis of 15 controlled clinical trials [22]. The authors reported that oral inositol supplementation reduced BMI and that this effect was particularly pronounced in people over 40 years of age. Younger age of patients participating in the present study, only mildly elevated mean values of BMI, the presence of a disorder predisposing to an increase in body weight, moderate doses of myo-inositol and metformin, and at most a moderate prolactin-lowering effect of the combination therapy explain why the changes in BMI did not reach the level of significance. However, the obtained results suggest that high-dose metformin/myo-inositol combination therapy may bring extra benefits if elevated prolactin levels are observed in obese subjects.

Owing to the lack of dedicated studies, the risk of prediabetes in individuals with hyperprolactinemia has not been calculated. However, unlike patients with prolactin levels within the reference range [23], human and animal studies indirectly suggest that supraphysiological prolactin concentrations may make patients more prone to developing impaired fasting glucose and/or impaired glucose tolerance [24-26]. Metabolic syndrome, which is commonly associated with prediabetes [27], has been diagnosed in 23–50% of patients with untreated or ineffectively treated hyperprolactinemia, and this risk was reduced to less than 5% in individuals receiving dopamine agonists for 6–60 months [24]. Moreover, high prolactin levels in premenopausal women were associated with hyperinsulinemia and insulin resistance [25]. Finally, animals with chronically elevated prolactin levels secondary to disruption of D2 receptors were characterized by glucose intolerance and impaired insulin response to glucose [26]. All these findings suggest that metformin/myo-inositol combination therapy, markedly improving glucose homeostasis, may more efficiently protect against diabetes than metformin alone, the risk of which is increased in the studied population [27, 28].

We can only speculate about the clinical significance of the impact on prolactin levels. Because the reduction in this hormone correlated with its baseline concentration, the impact of metformin/myo-inositol combination therapy on lactotroph secretory function is probably greatest in women with the highest hormone concentrations. This means that the combination therapy may be useful in women of reproductive age with resistant prolactinomas. Because individuals with prolactinoma were not included in the present study, this hypothesis should be verified in future studies. Metformin/myo-inositol combination therapy may also bring benefits to hyperprolactinemic women in whom dopamine agonists, the drugs of choice in the treatment of prolactin excess [28], are poorly tolerated. Finally, this combination therapy may be an interesting treatment option for patients with antipsychotic-induced hyperprolactinemia. The use of dopaminergic agonists in this group of patients is highly controversial given the risk of exacerbating psychotic symptoms and only limited effectiveness [29]. Apart from a reduction in prolactin levels, metformin/myo-inositol combination therapy can prevent deterioration of insulin sensitivity, which is often observed in patients receiving antipsychotics [30].

This research though is subject to several limitations. The study included a relatively small number of patients and was nonrandomized (a cohort study). All participants had prediabetes, and therefore it cannot be excluded that the effect on circulating prolactin levels may be different in individuals with normal glucose tolerance. The study included patients with hyperprolactinemia of various origins. Because tumor-induced hyperprolactinemia was one of the exclusion criteria, the question whether similar relationships between metformin and myo-inositol are observed in case of tumoral hyperprolactinemia requires further research. Macroprolactin content was determined using the polyethylene glycol precipitation method, although the gold standard for its measurement is gel filtration chromatography, which is, however, a time-consuming and expensive method [31]. Finally, it remains unanswered whether myo-inositol determines metformin action in men, not participating in the current study.

In conclusion, despite improving glucose homeostasis markers in both study groups, metformin resulted in a significant reduction in prolactin levels only in women concomitantly treated with myo-inositol. Treatment-induced changes in prolactin levels correlated with baseline prolactin levels and the impact of treatment on insulin sensitivity. The obtained results suggest that myo-inositol supplementation may potentiate the effect of metformin on lactotroph function. These preliminary findings need to be confirmed in randomized controlled trials.

Statement of Ethics

The study was conducted according to the guidelines of the Declaration of Helsinki and approved by the Institutional Review Board (the Bioethical Committee of the Medical University of Silesia [KNW/0022/KB/208/17–May 16, 2017]). All participants provided written informed consent prior to enrollment in the study.

Conflict of Interest Statement

The authors have no conflicts of interest to disclose.

Funding Sources

The study was supported by the statutory grant of the Medical University of Silesia (PCN-1-185/N/9/O).

Author Contributions

Robert Krysiak conceived the study, participated in its design, performed the statistical analysis, as well as drafted and edited the manuscript. Marcin Basiak conducted the literature search, carried out the assays, and performed the statistical analysis. Witold Szkróbka participated in the study design and provided critical input during manuscript preparations. Bogusław Okopień participated in the study design and coordination and provided critical input during manuscript preparations. All authors read and approved the final manuscript.

Data Availability Statement

All data generated or analyzed during this study are included in this manuscript. Further inquiries can be directed to the corresponding author.

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