Berberine Regulates GPX4 to Inhibit Ferroptosis of Islet β Cells

• Abstract
• Introduction
• Results and Discussion
• Materials and Methods
• Cell culture and transfection
• BBR treatment
• MTT assay
• Colony formation assay
• Iron content analysis
• ROS detection
• Western blot
• qRT-PCR
• Data analysis
• Contributorsʼ Statement
• References

Abstract

Ferroptosis, as a kind of non-apoptotic cell death, is involved in the pathogenesis of type 1 diabetes mellitus (T1DM). Islet B cells mainly produce insulin that is used to treat diabetes. Berberine (BBR) can ameliorate type 2 diabetes and insulin resistance in many ways. However, a few clues concerning the mechanism of BBR regulating ferroptosis of islet β cells in T1DM have been detected so far. We measured the effects of BBR and GPX4 on islet β cell viability and proliferation by MTT and colony formation assays. Western blot and qRT-PCR were utilized to examine GPX4 expression in islet β cells with distinct treatments. The influence of BBR and GPX4 on ferroptosis of islet β cells was investigated by evaluating the content of Fe²⁺ and reactive oxygen species (ROS) in cells. The mechanism of BBR targeting GPX4 to inhibit ferroptosis of islet β cells was further revealed by the rescue experiment. Our results showed that BBR and overexpression of GPX4 could notably accelerate cell viability and the proliferative abilities of islet β cells. Moreover, BBR stimulated GPX4 expression to reduce the content of Fe²⁺ and ROS, thereby repressing the ferroptosis of islet β cells, which functioned similarly as ferroptosis inhibitor Fer-1. In conclusion, BBR suppressed ferroptosis of islet β cells via promoting GPX4 expression, providing new insights into the mechanism of BBR for islet β cells.

Introduction

Diabetes mellitus (DM), as a hyperglycemia-characterized chronic metabolic disease, has become a major public health concern threatening human health [1]. In line with the statistics published by International Diabetes Federation [1], the number of diabetics (20 – 79 years old) in China has risen, and the incidence of diabetes is approaching 10%. China has gradually developed into the country with the largest number of diabetes population in the world [2]. The common DM subtypes consist of type 1 DM (T1DM) and type 2 DM (T2DM), with the former accounting for about 5% of all diabetics, whose incidence is increasing at a rate of 3 – 4% per year [3], [4]. T1DM is caused by the interaction of diverse genetic and environmental factors, and it occurs mostly among children aged 10 – 14, which has a serious influence on the health of adolescents [5], [6]. Studies have demonstrated that the pathogenesis of T1DM is due to insulin deficiency caused by the destruction of insulin-producing islet β cells by organ-specific immunity [7], [8]. Islet β cells maintain physiological glucose levels within a relatively constant range by sensing glucose and releasing insulin [9]. Once these cells are destroyed, blood glucose in T1DM patients is out of control, and even the use of insulin replacement therapy can lead to acute diseases (such as severe hypoglycemia and ketoacidosis) [10] and secondary complications (including blindness, heart disease, and renal failure) [11], [12]. Thus, it is helpful to accurately elucidate the mechanism of β cell immune destruction and explore therapeutic drugs for developing new therapeutic strategies, which are greatly significant to control and ameliorate T1DM.

Natural drugs, especially Chinese herbal medicines, can regulate body functions through multiple targets and mechanisms and have been proven to have mild and notable effects on reducing blood sugar and blood pressure [13], [14], [15]. There are more than 80 kinds of traditional Chinese medicine commonly used for diabetes and its complications. Berberine (BBR) has aroused our research interest due to its favorable hypoglycemic activity [16]. A previous study exhibited that BBR can promote SIRT3 ubiquitination to enhance glucose uptake in ovarian cells and hinder mitochondrial function, which can improve insulin sensitivity in patients with polycystic ovary syndrome [17]. Among DM-related studies, Qin et al. [18] found that BBR prevents diabetic kidney disease by accelerating PGc-1α-regulated mitochondrial energy homeostasis, using DKD mouse models. Additionally, BBR can relieve T2DM symptoms via changing intestinal microbiota and reducing aromatic amino acids [19], but its specific molecular mechanism affecting T1DM requires further exploration.

Ferroptosis is a newly characterized iron-dependent non-apoptotic form of cell death caused by the imbalance of the generation and degradation of intracellular lipid reactive oxygen species (ROS) [20]. Ferroptosis is rigorously controlled by intracellular signaling pathways. The main function of glutathione-dependent antioxidant oxidase 4 (GPX4) is to eliminate excess ROS that induces DNA damage. The abnormal GPX4 expression leads to the reduced antioxidant capacity of cells, ROS accumulation, and ultimately ferroptosis of cells [21]. The latest study indicated that ferroptosis can generate the loss and dysfunction of pancreatic β cells. Inhibiting the accumulation of Fe²⁺ and ferroptosis of pancreatic β cells contributes to the disease control of T2DM mice [22]. Nevertheless, the relationship between T1DM pathogenesis and ferroptosis of islet β cells needs to be further investigated.

Here, we observed that BBR arrested ferroptosis of islet β cells in vitro. Moreover, BBR could enhance GPX4 expression, and hinder the accumulation of Fe²⁺ content and ROS production in islet β cells. Our findings may offer novel insights into the mechanism by which BBR treating T1DM.I

Results and Discussion

We initially investigated the influence of BBR on the proliferation of islet β cells. Cell viability and colony formation assays confirmed that 10 – 40 µM BBR promoted the proliferation of islet β cells NIT-1 in a dose-dependent manner ([Fig. 1 a, b]). Erastin is a frequent inducer of ferroptosis that can bind to and arrest voltage-dependent anionic pathways (VDAC2/VDAC3), and Fer-1 can suppress Erastin [23], [24]. In this study, we discovered that BBR acts similarly to ferroptosis inhibitor Fer-1 in reversing Erastin-induced decline in cell viability and colony formation ([Fig. 1 c, d]). The main manifestations of ferroptosis are ROS increase, Fe²⁺ accumulation, and the expression change of GPX4 and other related proteins [25], [26], [27]. To clarify whether the promotion of BBR on the proliferation of islet β cells was related to ferroptosis inhibition, we further examined ROS level, Fe²⁺ accumulation, and the expression of ferroptosis-related protein GXP4 in islet β cells under BBR treatment. The results indicated that BBR, like Fer-1, could reduce Fe²⁺ accumulation and ROS level in islet β cells ([Fig. 1 e, f]). In addition, BBR may enhance the expression of GXP4 protein and mRNA in islet β cells ([Fig. 1 g, h]). Overall, BBR could arrest ferroptosis of islet β cells. RSL3 and Erastin have the same effect, and both are iron death agonists. We also applied RSL3 in our experiment to study the effect of RSL3 on GPX4. The results showed that RSL3 inhibited the proliferation and clonal function of islet β-cells, and promoted the ferroptosis of islet β-cells. In addition, BBR reversed the above-mentioned inhibitory effect and promoting effect of RSL3 on the phenotypes of islet β-cells (Fig. 1S, Supporting Information).

A study revealed that ferroptosis is a new form of islet β cell death, and GPX4 is a key molecule involved in regulating ferroptosis [28]. To analyze the effect of GPX4 on ferroptosis of islet β-cells, we constructed islet β cells with GPX4 overexpression or knockdown. First, validation of GPX4 transfection efficiency was performed using qRT-PCR ([Fig. 2 a]). Subsequently, we investigated that GPX4 overexpression could ameliorate the cell viability and colony formation ability of islet β cells NIT-1, while GPX4 knockdown evidently restrained these abilities ([Fig. 2 b, c]). Fe²⁺ content analysis and ROS level detection illustrated that GPX4 overexpression inhibited Fe²⁺ accumulation in NIT-1 cells, and the intracellular ROS level decreased notably; GPX4 knockdown accelerated Fe2+ accumulation in NIT-1 cells, and the intracellular ROS level increased remarkably ([Fig. 2 d, e]) These data suggested that GPX4 upregulation markedly restrained ferroptosis of islet β cells.

In previous studies, BBR was discovered to enhance the viability of islet β cells and upregulate intracellular GPX4 protein expression. Hence, we hypothesized that BBR targeted GPX4 to arrest ferroptosis of islet β cells, which was further verified by the rescue experiment. The addition of 10 µM BBR to GPX4-inhibited islet β cells NIT-1 significantly restored cell viability and colony formation abilities ([Fig. 3 a, b]). Subsequently, we detected ferroptosis-related protein expression, Fe²⁺ content, and ROS content in cells, and the results demonstrated that the addition of BBR partially restored GPX4 expression and intracellular Fe²⁺ content in islet β cells with GPX4 inhibition ([Fig. 3 c, d, e]), and also reduced ROS level ([Fig. 3 f]). These data hinted that BBR targeting GPX4 led to ferroptosis inhibition in islet β cells.

T1DM is a chronic autoimmune disease in young patients and is characterized by loss of islet β cells [29]. Currently, exogenous insulin injections are the main life-sustaining treatment for millions of patients. Nonetheless, insulin injection is incapable of effectively controlling the underlying autoimmunity, nor reversing β cell destruction or aiding cell regeneration [30]. Moreover, insulin treatment can cause weight gain and increase the risk of hypoglycemia. Thus, there is an urgent need for therapies that avoid exogenous insulin administration in DM patients. As a classical natural medicine, BBR has been proven to function profoundly in the treatment of cardiovascular metabolic diseases such as hyperglycemia and hypertension [31], [32]. Muhammad et al. [33] proposed BBR efficacy in the treatment of T2DM patients and explored that BBR administration (three times a day at 500 mg each dose) reduces serum pyruvate aldehyde level and insulin resistance through improving blood glucose control. In addition, among related studies on T1DM treatment, Chueh et al. [34], [35] suggested that fasting glucose level decreases in a dose-dependent manner and insulin levels increase in non-obese diabetic mice (NOD) with spontaneous T1DM after 14 weeks of oral BBR. Cui et al. [36] also obtained consistent results by the construction of animal model and observed that oral BBR (200 mg/kg) for two weeks can effectively prevent T1DM progression in NOD mice and reduce the secretion of cytokines Th17 and Th1. It thus follows that BBR played a potential therapeutic role in T1DM, but its specific mechanism has not been clearly explored.

Herein, we investigated that treatment with 10 µM BBR could not only stimulate the proliferative ability of islet β cells NIT-1 but also reduce the content of Fe²⁺ and ROS, which was similar to Fer-1. Fer-1 exerts a negative modulatory effect on the occurrence of ferroptosis. Byoung-Seob et al. [37] elucidated that BBR enhances insulin secretion and proliferation in glucose-stimulated NIT-1 cells through ameliorating the insulin/insulin-like growth factor-1 signaling cascade. Another study showed that BBR had an effect on cell proliferation/viability and ferroptotic markers after 12 weeks of mice feeding [38]. However, the duration of our experiments was not long enough to sustain the effect of BBR on cell proliferation/viability and ferroptotic markers, the reliability of our experimental results was therefore not affected. Further cell experiments exhibited that ferroptosis-related protein GPX4 could accelerate cell viability and colony formation abilities of islet β cells NIT-1 and reduce ferroptosis levels. This is in line with the results of Bastian et al. [39], which proposed that GPX4 overexpression effectively alleviates tert-BHP-induced islet β cell death in mice. Subsequent rescue experiments further illustrated that BBR could partially restore GPX4 expression in GPX4-knocked down islet β cells, restored cell proliferative ability and reduced intracellular Fe²⁺ and ROS levels, suggesting that BBR hindered ferroptosis of islet β cells via targeting GPX4. It was further elucidated that BBR had the potential as a drug for the treatment of T1DM. Xiaoyan Lv et al. [40] showed that BBR treatment effectively improved insulin synthesis, reduced miR-204 level, and increased SIRT1 expression in pancreatic islets of diabetic mice. However, due to the limited experimental conditions, we did not conduct in vivo mouse model construction and BBR treatment experiments, which were the limitations of our study. We will extend the relevant content in the future.

Beginning with BBR and T1DM, we investigated the mechanism of BBR modulating ferroptosis of islet β cells, in other words, BBR inhibited ferroptosis of islet β cells by targeting GPX4. However, despite these advances, this study lacks further verification of in vivo experiments. In the future, the regulation of BBR on GPX4 gene and the effect on ferroptosis of islet β cells in vivo will be verified by constructing mouse models.

Materials and Methods

Cell culture and transfection

Mice insulinoma β cells NIT-1 (BNCC340516) were procured from BeNa Culture Collection (BNCC). These cells were cultured in high glucose-Dulbeccoʼs Modified Eagle Medium (DMEM, Thermo Fisher Scientific) containing 10% fetal bovine serum (FBS), and the medium was kept in an incubator at 37 °C with 5% CO₂ (Thermo Fisher Scientific). NIT-1 cells were transfected with GPX4 knockdown lentivirus (LV-2 vector) or overexpression (LV5 vector), and blank control lentivirus vector LC (GenePharma) and underwent 20 purification screenings until a cell line that could be stably expressed emerges.

BBR treatment

BBR was purchased from Abcam Company (#ab142117) and dissolved in ultra-pure water with a concentration of 5 mM of mother liquor. BBR mother liquor was diluted to the required concentration with fresh medium prior to use. In this study, NIT-1 cells were treated with 5, 10, 20, and 40 µM of BBR for 24 h, and the lowest BBR concentration (10 µM) which could remarkably facilitate cell proliferation was selected as the required concentration for subsequent experiments.

MTT assay

NIT-1 cells were seeded into 96-well plates (8 × 10³ cells/well) and incubated for 24 h. 10 mg/mL MTT solution (Merck & Co.,) was applied to treat the cells for 4 h. The medium was thenremoved and 150 µL Dimethyl sulfoxide (DMSO) was added to incubate cells for 10 min in a 37 °C incubator. Finally, the optical density (OD) of each well was observed at 570 nm by using a microplate reader.

Colony formation assay

Cells were suspended by 0.25% trypsin (Thermo Fisher Scientific) and then plated into 6 cm petri dish with 800 cells in each dish. Cells cultured in high-glucose DMEM with 10% FBS were maintained with 5% CO₂ and 37 °C for 2 – 3 weeks. The medium was removed when colonies were formed visibly, and the colonies were carefully rinsed with phosphate-buffered saline (PBS). Cells were fixed with 4% paraformaldehyde for 15 min, and dyed with 0.1% crystal violet for 20 min. The excess crystal violet dye in the wells was washed with PBS. Finally, the images were taken and the number of colonies in the wells was counted.

Iron content analysis

In accordance with the manufacturerʼs instructions, Fe²⁺ and total iron content in the cells were determined by the iron detection kit (Sigma-Aldrich). First, 2 × 10⁶ cells were suspended with trypsin and added with iron detection buffer. Cells were centrifuged, followed by the transfer of the collected supernatant to 96-well plates with 100 µl per well. 5 µl iron detection buffer was then added to each well, and the samples were mixed and incubated at room temperature without light for 30 min. 100 µl iron probe was supplemented to each well, and the samples were incubated in the dark for 60 min at room temperature. Lastly, the OD value was measured at 593 nm via a microplate reader.

ROS detection

Cells were cultured in a 6 well plate with 1 mL/well DCFH-DA probe (diluted 1 : 1000, Beyotime, China) at 37 °C for 25 min. Afterward, cells were washed with PBS twice, and a confocal microscopy (Thermo Fisher Scientific) was applied to detect fluorescence and photograph cells. Fluorescence intensity was quantified by ImageJ software.

Western blot

The cells were lysed with protein lysate (Thermo Fisher Scientific) and protein concentration was determined by a bicinchoninic acid protein detection kit (Thermo Fisher Scientific) after ultrasonic fragmentation. Protein samples with equal quantity were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. After the protein was transferred to a polyvinylidene fluoride membrane, the membrane was blocked with 3% albumin from bovine serum at room temperature for 1 h. The membrane containing the target band was then incubated with primary antibodies (1 : 1000, ab41787, Abcam, UK) at 4 °C overnight. The next day, the membrane was rinsed with Tris-Buffered Saline Tween-20 (TBST) and cultured with horseradish peroxidase-conjugated secondary antibody (1 : 2000, ab6721, Abcam, UK) in a shaker for 2 h at room temperature. The expression of proteins on the membrane was developed utilizing enhanced chemiluminescence kit (Thermo Fisher Scientific).Both primary antibodies and the secondary antibody were provided by Cell Signaling Technology which were anti-GPX4 (#52455, rabbit-derived), anti-GAPDH (#5174, rabbit-derived), and anti-rabbit IgG (#14708) antibodies, respectively.

qRT-PCR

Total RNA separated from harvested cells (10⁶) were isolated adopting Trizol reagent (Thermo Fisher Scientific). The extracted RNA was reversely transcribed into cDNA employing PrimeScript RT kit (Takara), and real-time PCR was carried out using SYBR Premix Ex TaqII (Takara). PCR was conducted at 94 °C for 3 min, 94 °C for 30 s, 54 °C for 30 s, 70 °C for 30 s, with 25 – 35 cycles, and finally extended at 72 °C for 10 min. GAPDH served as the internal reference, and the fold-change was calculated by 2^-△△CT. The primer sequences are as follows:

GAPDH-F, 5′-GGACCTGACCTGCCGTCTAG-3′;

GAPDH-R, 5′-GTAGCCCAGGATGCCCTTGA-3′;

GPX4-F, 5′-ATACGCTGAGTGTGGTTTGC-3′;

GPX4-R, 5′-CTTCATCCACTTCCACAGCG-3′.

Data analysis

Data were displayed as mean ± standard deviation. Each assay was repeated 3 times. Intergroup differences were analyzed by Studentʼs t-test or one-way analysis of variance. GraphPad Prism 8 software (GraphPad Software) was employed for analysis, and p < 0.05 was thought to be statistically significant.

Contributorsʼ Statement

Lei Bao: conceptualization, methodology, writing – original draft; Yixuan Jin: conceptualization, data curation, writing – review & editing; Jiani Han: validation, investigation; Wanqiu Wang: formal analysis, data curation; Lingling Qian: resources, visualization; Weiming Wu: project administration, writing – original draft.

Conflict of Interest

The authors declare that they have no conflict of interest.

• References

• 1 Ogurtsova K, da Rocha Fernandes JD, Huang Y, Linnenkamp U, Guariguata L, Cho NH, Cavan D, Shaw JE, Makaroff LE. IDF Diabetes Atlas: Global estimates for the prevalence of diabetes for 2015 and 2040. Diabetes Res Clin Pract 2017; 128: 40-50
  CrossrefPubMedSearch in Google Scholar
• 2 Guariguata L, Whiting DR, Hambleton I, Beagley J, Linnenkamp U, Shaw JE. Global estimates of diabetes prevalence for 2013 and projections for 2035. Diabetes Res Clin Pract 2014; 103: 137-149
  CrossrefPubMedSearch in Google Scholar
• 3 Norris JM, Johnson RK, Stene LC. Type 1 diabetes-early life origins and changing epidemiology. Lancet Diabetes Endocrinol 2020; 8: 226-238
  Search in Google Scholar
• 4 American Diabetes Association. (2) Classification and diagnosis of diabetes. Diabetes Care 2015; 38: S8-S16
  CrossrefPubMedSearch in Google Scholar
• 5 Weng J, Zhou Z, Guo L, Zhu D, Ji L, Luo X, Mu Y, Jia W. T1D China Study Group. Incidence of type 1 diabetes in China, 2010–13: Population based study. BMJ 2018; 360: j5295
  CrossrefPubMedSearch in Google Scholar
• 6 Cerna M. Epigenetic regulation in etiology of type 1 diabetes mellitus. Int J Mol Sci 2019; 21: 36
  CrossrefPubMedSearch in Google Scholar
• 7 Haak T, Gölz S, Fritsche A, Füchtenbusch M, Siegmund T, Schnellbächer E, Klein HH, Uebel T, Droßel D. Therapy of type 1 diabetes. Exp Clin Endocrinol Diabetes 2019; 127: S27-S38
  Thieme ConnectPubMedSearch in Google Scholar
• 8 Anderson MS, Bluestone JA. The NOD mouse: A model of immune dysregulation. Annu Rev Immunol 2005; 23: 447-485
  CrossrefPubMedSearch in Google Scholar
• 9 Bluestone JA, Herold K, Eisenbarth G. Genetics, pathogenesis and clinical interventions in type 1 diabetes. Nature 2010; 464: 1293-1300
  CrossrefPubMedSearch in Google Scholar
• 10 Vauzelle-Kervroedan F, Delcourt C, Forhan A, Jougla E, Hatton F, Papoz L. Analysis of mortality in French diabetic patients from death certificates: A comparative study. Diabetes Metab 1999; 25: 404-411
  PubMedSearch in Google Scholar
• 11 Writing Team for the Diabetes Control and Complications Trial/Epidemiology of Diabetes Interventions and Complications Research Group. Sustained effect of intensive treatment of type 1 diabetes mellitus on development and progression of diabetic nephropathy: The Epidemiology of Diabetes Interventions and Complications (EDIC) study. JAMA 2003; 290: 2159-2167
  CrossrefPubMedSearch in Google Scholar
• 12 Maahs DM, Rewers M. Editorial: Mortality and renal disease in type 1 diabetes mellitus–progress made, more to be done. J Clin Endocrinol Metab 2006; 91: 3757-3759
  CrossrefPubMedSearch in Google Scholar
• 13 Pang B, Zhao LH, Zhou Q, Zhao TY, Wang H, Gu CJ, Tong XL. Application of berberine on treating type 2 diabetes mellitus. Int J Endocrinol 2015; 2015: 905749
  CrossrefPubMedSearch in Google Scholar
• 14 Li WL, Zheng HC, Bukuru J, De Kimpe N. Natural medicines used in the traditional Chinese medical system for therapy of diabetes mellitus. J Ethnopharmacol 2004; 92: 1-21
  CrossrefPubMedSearch in Google Scholar
• 15 El-Kaissi S, Sherbeeni S. Pharmacological management of type 2 diabetes mellitus: An update. Curr Diabetes Rev 2011; 7: 392-405
  CrossrefPubMedSearch in Google Scholar
• 16 Baska A, Leis K, Galazka P. Berberine in the treatment of diabetes mellitus: A review. Endocr Metab Immune Disord Drug Targets 2021; 21: 1379-1386
  CrossrefPubMedSearch in Google Scholar
• 17 Li W, Li D, Kuang H, Feng X, Ai W, Wang Y, Shi S, Chen J, Fan R. Berberine increases glucose uptake and intracellular ROS levels by promoting Sirtuin 3 ubiquitination. Biomed Pharmacother 2020; 121: 109563
  CrossrefPubMedSearch in Google Scholar
• 18 Qin X, Jiang M, Zhao Y, Gong J, Su H, Yuan F, Fang K, Yuan X, Yu X, Dong H, Lu F. Berberine protects against diabetic kidney disease via promoting PGC-1alpha-regulated mitochondrial energy homeostasis. Br J Pharmacol 2020; 177: 3646-3661
  CrossrefPubMedSearch in Google Scholar
• 19 Yao Y, Chen H, Yan L, Wang W, Wang D. Berberine alleviates type 2 diabetic symptoms by altering gut microbiota and reducing aromatic amino acids. Biomed Pharmacother 2020; 131: 110669
  CrossrefPubMedSearch in Google Scholar
• 20 Hou Y, Cai S, Yu S, Lin H. Metformin induces ferroptosis by targeting miR-324-3p/GPX4 axis in breast cancer. Acta Biochim Biophys Sin (Shanghai) 2021; 53: 333-341
  CrossrefPubMedSearch in Google Scholar
• 21 Zhou B, Liu J, Kang R, Klionsky DJ, Kroemer G, Tang D. Ferroptosis is a type of autophagy-dependent cell death. Semin Cancer Biol 2020; 66: 89-100
  CrossrefPubMedSearch in Google Scholar
• 22 Liu J, Cheng C, Jiao J, Huang W, Huang J, Sun J, Shao J. MircoRNA-129-5p suppresses the development of glioma by targeting HOXC10. Pathol Res Pract 2020; 216: 152868
  CrossrefPubMedSearch in Google Scholar
• 23 Geng N, Shi BJ, Li SL, Zhong ZY, Li YC, Xua WL, Zhou H, Cai JH. Knockdown of ferroportin accelerates erastin-induced ferroptosis in neuroblastoma cells. Eur Rev Med Pharmacol Sci 2018; 22: 3826-3836
  PubMedSearch in Google Scholar
• 24 Shintoku R, Takigawa Y, Yamada K, Kubota C, Yoshimoto Y, Takeuchi T, Koshiishi I, Torii S. Lipoxygenase-mediated generation of lipid peroxides enhances ferroptosis induced by erastin and RSL3. Cancer Sci 2017; 108: 2187-2194
  CrossrefPubMedSearch in Google Scholar
• 25 Lu B, Chen XB, Hong YC, Zhu H, He QJ, Yang B, Ying MD, Cao J. Identification of PRDX6 as a regulator of ferroptosis. Acta Pharmacol Sin 2019; 40: 1334-1342
  CrossrefPubMedSearch in Google Scholar
• 26 Dixon SJ, Lemberg KM, Lamprecht MR, Skouta R, Zaitsev EM, Gleason CE, Patel DN, Bauer AJ, Cantley AM, Yang WS, Morrison 3rd B, Stockwell BR. Ferroptosis: An iron-dependent form of nonapoptotic cell death. Cell 2012; 149: 1060-1072
  CrossrefPubMedSearch in Google Scholar
• 27 Liu P, Feng Y, Li H, Chen X, Wang G, Xu S, Li Y, Zhao L. Ferrostatin-1 alleviates lipopolysaccharide-induced acute lung injury via inhibiting ferroptosis. Cell Mol Biol Lett 2020; 25: 10
  CrossrefPubMedSearch in Google Scholar
• 28 Li D, Jiang C, Mei G, Zhao Y, Chen L, Liu J, Tang Y, Gao C, Yao P. Quercetin alleviates ferroptosis of pancreatic β cells in type 2 diabetes. Nutrients 2020; 12: 2954
  CrossrefPubMedSearch in Google Scholar
• 29 El-Mokhtar MA, Elsherbiny NM, Sayed D, Raafat DM, Askar E, Hussein A, Abdel-Malek MAY, Shalaby AM. Altered regulatory B cell subsets in children with type 1 diabetes mellitus. J Immunol Res 2020; 2020: 8935694
  PubMedSearch in Google Scholar
• 30 Stiner R, Alexander M, Liu G, Liao W, Liu Y, Yu J, Pone EJ, Zhao W, Lakey JRT. Transplantation of stem cells from umbilical cord blood as therapy for type I diabetes. Cell Tissue Res 2019; 378: 155-162
  CrossrefPubMedSearch in Google Scholar
• 31 Suadoni MT, Atherton I. Berberine for the treatment of hypertension: A systematic review. Complement Ther Clin Pract 2021; 42: 101287
  CrossrefPubMedSearch in Google Scholar
• 32 Feng X, Sureda A, Jafari S, Memariani Z, Tewari D, Annunziata G, Barrea L, Hassan STS, Šmejkal K, Malaník M, Sychrová A, Barreca D, Ziberna L, Mahomoodally MF, Zengin G, Xu S, Nabavi SM, Shen AZ. Berberine in cardiovascular and metabolic diseases: From mechanisms to therapeutics. Theranostics 2019; 9: 1923-1951
  CrossrefPubMedSearch in Google Scholar
• 33 Memon MA, Khan RN, Riaz S, Ain QU, Ahmed M, Kumar N. Methylglyoxal and insulin resistance in berberine-treated type 2 diabetic patients. J Res Med Sci 2018; 23: 110
  CrossrefPubMedSearch in Google Scholar
• 34 Chueh WH, Lin JY. Berberine, an isoquinoline alkaloid in herbal plants, protects pancreatic islets and serum lipids in nonobese diabetic mice. J Agric Food Chem 2011; 59: 8021-8027
  CrossrefPubMedSearch in Google Scholar
• 35 Chueh WH, Lin JY. Protective effect of berberine on serum glucose levels in non-obese diabetic mice. Int Immunopharmacol 2012; 12: 534-538
  CrossrefPubMedSearch in Google Scholar
• 36 Cui G, Qin X, Zhang Y, Gong Z, Ge B, Zang YQ. Berberine differentially modulates the activities of ERK, p38 MAPK, and JNK to suppress Th17 and Th1 T cell differentiation in type 1 diabetic mice. J Biol Chem 2009; 284: 28420-28429
  CrossrefPubMedSearch in Google Scholar
• 37 Ko BS, Choi SB, Park SK, Jang JS, Kim YE, Park S. Insulin sensitizing and insulinotropic action of berberine from Cortidis rhizoma. Biol Pharm Bull 2005; 28: 1431-1437
  CrossrefPubMedSearch in Google Scholar
• 38 Wu YS, Li ZM, Chen YT, Dai SJ, Zhou XJ, Yang YX, Lou JS, Ji LT, Bao YT, Xuan L, Lin LN, Li CY. Berberine improves inflammatory responses of diabetes mellitus in zucker diabetic fatty rats and insulin-resistant HepG2 cells through the PPM1B pathway. J Immunol Res 2020; 2020: 2141508
  PubMedSearch in Google Scholar
• 39 Krümmel B, Plötz T, Jörns A, Lenzen S, Mehmeti I. The central role of glutathione peroxidase 4 in the regulation of ferroptosis and its implications for pro-inflammatory cytokine-mediated beta-cell death. Biochim Biophys Acta Mol Basis Dis 2021; 1867: 166114
  CrossrefPubMedSearch in Google Scholar
• 40 Lv X, Zhao Y, Yang X, Han H, Ge Y, Zhang M, Zhang H, Zhang M, Chen L. Berberine potentiates insulin secretion and prevents β-cell dysfunction through the miR-204/SIRT1 signaling pathway. Front Pharmacol 2021; 12: 720866
  CrossrefPubMedSearch in Google Scholar

Correspondence

Publication History

Received: 25 February 2022

Accepted after revision: 15 August 2022

Article published online:
09 November 2022

© 2022. Thieme. All rights reserved.

Georg Thieme Verlag KG
Rüdigerstraße 14, 70469 Stuttgart, Germany

• References

• 1 Ogurtsova K, da Rocha Fernandes JD, Huang Y, Linnenkamp U, Guariguata L, Cho NH, Cavan D, Shaw JE, Makaroff LE. IDF Diabetes Atlas: Global estimates for the prevalence of diabetes for 2015 and 2040. Diabetes Res Clin Pract 2017; 128: 40-50
  CrossrefPubMedSearch in Google Scholar
• 2 Guariguata L, Whiting DR, Hambleton I, Beagley J, Linnenkamp U, Shaw JE. Global estimates of diabetes prevalence for 2013 and projections for 2035. Diabetes Res Clin Pract 2014; 103: 137-149
  CrossrefPubMedSearch in Google Scholar
• 3 Norris JM, Johnson RK, Stene LC. Type 1 diabetes-early life origins and changing epidemiology. Lancet Diabetes Endocrinol 2020; 8: 226-238
  Search in Google Scholar
• 4 American Diabetes Association. (2) Classification and diagnosis of diabetes. Diabetes Care 2015; 38: S8-S16
  CrossrefPubMedSearch in Google Scholar
• 5 Weng J, Zhou Z, Guo L, Zhu D, Ji L, Luo X, Mu Y, Jia W. T1D China Study Group. Incidence of type 1 diabetes in China, 2010–13: Population based study. BMJ 2018; 360: j5295
  CrossrefPubMedSearch in Google Scholar
• 6 Cerna M. Epigenetic regulation in etiology of type 1 diabetes mellitus. Int J Mol Sci 2019; 21: 36
  CrossrefPubMedSearch in Google Scholar
• 7 Haak T, Gölz S, Fritsche A, Füchtenbusch M, Siegmund T, Schnellbächer E, Klein HH, Uebel T, Droßel D. Therapy of type 1 diabetes. Exp Clin Endocrinol Diabetes 2019; 127: S27-S38
  Thieme ConnectPubMedSearch in Google Scholar
• 8 Anderson MS, Bluestone JA. The NOD mouse: A model of immune dysregulation. Annu Rev Immunol 2005; 23: 447-485
  CrossrefPubMedSearch in Google Scholar
• 9 Bluestone JA, Herold K, Eisenbarth G. Genetics, pathogenesis and clinical interventions in type 1 diabetes. Nature 2010; 464: 1293-1300
  CrossrefPubMedSearch in Google Scholar
• 10 Vauzelle-Kervroedan F, Delcourt C, Forhan A, Jougla E, Hatton F, Papoz L. Analysis of mortality in French diabetic patients from death certificates: A comparative study. Diabetes Metab 1999; 25: 404-411
  PubMedSearch in Google Scholar
• 11 Writing Team for the Diabetes Control and Complications Trial/Epidemiology of Diabetes Interventions and Complications Research Group. Sustained effect of intensive treatment of type 1 diabetes mellitus on development and progression of diabetic nephropathy: The Epidemiology of Diabetes Interventions and Complications (EDIC) study. JAMA 2003; 290: 2159-2167
  CrossrefPubMedSearch in Google Scholar
• 12 Maahs DM, Rewers M. Editorial: Mortality and renal disease in type 1 diabetes mellitus–progress made, more to be done. J Clin Endocrinol Metab 2006; 91: 3757-3759
  CrossrefPubMedSearch in Google Scholar
• 13 Pang B, Zhao LH, Zhou Q, Zhao TY, Wang H, Gu CJ, Tong XL. Application of berberine on treating type 2 diabetes mellitus. Int J Endocrinol 2015; 2015: 905749
  CrossrefPubMedSearch in Google Scholar
• 14 Li WL, Zheng HC, Bukuru J, De Kimpe N. Natural medicines used in the traditional Chinese medical system for therapy of diabetes mellitus. J Ethnopharmacol 2004; 92: 1-21
  CrossrefPubMedSearch in Google Scholar
• 15 El-Kaissi S, Sherbeeni S. Pharmacological management of type 2 diabetes mellitus: An update. Curr Diabetes Rev 2011; 7: 392-405
  CrossrefPubMedSearch in Google Scholar
• 16 Baska A, Leis K, Galazka P. Berberine in the treatment of diabetes mellitus: A review. Endocr Metab Immune Disord Drug Targets 2021; 21: 1379-1386
  CrossrefPubMedSearch in Google Scholar
• 17 Li W, Li D, Kuang H, Feng X, Ai W, Wang Y, Shi S, Chen J, Fan R. Berberine increases glucose uptake and intracellular ROS levels by promoting Sirtuin 3 ubiquitination. Biomed Pharmacother 2020; 121: 109563
  CrossrefPubMedSearch in Google Scholar
• 18 Qin X, Jiang M, Zhao Y, Gong J, Su H, Yuan F, Fang K, Yuan X, Yu X, Dong H, Lu F. Berberine protects against diabetic kidney disease via promoting PGC-1alpha-regulated mitochondrial energy homeostasis. Br J Pharmacol 2020; 177: 3646-3661
  CrossrefPubMedSearch in Google Scholar
• 19 Yao Y, Chen H, Yan L, Wang W, Wang D. Berberine alleviates type 2 diabetic symptoms by altering gut microbiota and reducing aromatic amino acids. Biomed Pharmacother 2020; 131: 110669
  CrossrefPubMedSearch in Google Scholar
• 20 Hou Y, Cai S, Yu S, Lin H. Metformin induces ferroptosis by targeting miR-324-3p/GPX4 axis in breast cancer. Acta Biochim Biophys Sin (Shanghai) 2021; 53: 333-341
  CrossrefPubMedSearch in Google Scholar
• 21 Zhou B, Liu J, Kang R, Klionsky DJ, Kroemer G, Tang D. Ferroptosis is a type of autophagy-dependent cell death. Semin Cancer Biol 2020; 66: 89-100
  CrossrefPubMedSearch in Google Scholar
• 22 Liu J, Cheng C, Jiao J, Huang W, Huang J, Sun J, Shao J. MircoRNA-129-5p suppresses the development of glioma by targeting HOXC10. Pathol Res Pract 2020; 216: 152868
  CrossrefPubMedSearch in Google Scholar
• 23 Geng N, Shi BJ, Li SL, Zhong ZY, Li YC, Xua WL, Zhou H, Cai JH. Knockdown of ferroportin accelerates erastin-induced ferroptosis in neuroblastoma cells. Eur Rev Med Pharmacol Sci 2018; 22: 3826-3836
  PubMedSearch in Google Scholar
• 24 Shintoku R, Takigawa Y, Yamada K, Kubota C, Yoshimoto Y, Takeuchi T, Koshiishi I, Torii S. Lipoxygenase-mediated generation of lipid peroxides enhances ferroptosis induced by erastin and RSL3. Cancer Sci 2017; 108: 2187-2194
  CrossrefPubMedSearch in Google Scholar
• 25 Lu B, Chen XB, Hong YC, Zhu H, He QJ, Yang B, Ying MD, Cao J. Identification of PRDX6 as a regulator of ferroptosis. Acta Pharmacol Sin 2019; 40: 1334-1342
  CrossrefPubMedSearch in Google Scholar
• 26 Dixon SJ, Lemberg KM, Lamprecht MR, Skouta R, Zaitsev EM, Gleason CE, Patel DN, Bauer AJ, Cantley AM, Yang WS, Morrison 3rd B, Stockwell BR. Ferroptosis: An iron-dependent form of nonapoptotic cell death. Cell 2012; 149: 1060-1072
  CrossrefPubMedSearch in Google Scholar
• 27 Liu P, Feng Y, Li H, Chen X, Wang G, Xu S, Li Y, Zhao L. Ferrostatin-1 alleviates lipopolysaccharide-induced acute lung injury via inhibiting ferroptosis. Cell Mol Biol Lett 2020; 25: 10
  CrossrefPubMedSearch in Google Scholar
• 28 Li D, Jiang C, Mei G, Zhao Y, Chen L, Liu J, Tang Y, Gao C, Yao P. Quercetin alleviates ferroptosis of pancreatic β cells in type 2 diabetes. Nutrients 2020; 12: 2954
  CrossrefPubMedSearch in Google Scholar
• 29 El-Mokhtar MA, Elsherbiny NM, Sayed D, Raafat DM, Askar E, Hussein A, Abdel-Malek MAY, Shalaby AM. Altered regulatory B cell subsets in children with type 1 diabetes mellitus. J Immunol Res 2020; 2020: 8935694
  PubMedSearch in Google Scholar
• 30 Stiner R, Alexander M, Liu G, Liao W, Liu Y, Yu J, Pone EJ, Zhao W, Lakey JRT. Transplantation of stem cells from umbilical cord blood as therapy for type I diabetes. Cell Tissue Res 2019; 378: 155-162
  CrossrefPubMedSearch in Google Scholar
• 31 Suadoni MT, Atherton I. Berberine for the treatment of hypertension: A systematic review. Complement Ther Clin Pract 2021; 42: 101287
  CrossrefPubMedSearch in Google Scholar
• 32 Feng X, Sureda A, Jafari S, Memariani Z, Tewari D, Annunziata G, Barrea L, Hassan STS, Šmejkal K, Malaník M, Sychrová A, Barreca D, Ziberna L, Mahomoodally MF, Zengin G, Xu S, Nabavi SM, Shen AZ. Berberine in cardiovascular and metabolic diseases: From mechanisms to therapeutics. Theranostics 2019; 9: 1923-1951
  CrossrefPubMedSearch in Google Scholar
• 33 Memon MA, Khan RN, Riaz S, Ain QU, Ahmed M, Kumar N. Methylglyoxal and insulin resistance in berberine-treated type 2 diabetic patients. J Res Med Sci 2018; 23: 110
  CrossrefPubMedSearch in Google Scholar
• 34 Chueh WH, Lin JY. Berberine, an isoquinoline alkaloid in herbal plants, protects pancreatic islets and serum lipids in nonobese diabetic mice. J Agric Food Chem 2011; 59: 8021-8027
  CrossrefPubMedSearch in Google Scholar
• 35 Chueh WH, Lin JY. Protective effect of berberine on serum glucose levels in non-obese diabetic mice. Int Immunopharmacol 2012; 12: 534-538
  CrossrefPubMedSearch in Google Scholar
• 36 Cui G, Qin X, Zhang Y, Gong Z, Ge B, Zang YQ. Berberine differentially modulates the activities of ERK, p38 MAPK, and JNK to suppress Th17 and Th1 T cell differentiation in type 1 diabetic mice. J Biol Chem 2009; 284: 28420-28429
  CrossrefPubMedSearch in Google Scholar
• 37 Ko BS, Choi SB, Park SK, Jang JS, Kim YE, Park S. Insulin sensitizing and insulinotropic action of berberine from Cortidis rhizoma. Biol Pharm Bull 2005; 28: 1431-1437
  CrossrefPubMedSearch in Google Scholar
• 38 Wu YS, Li ZM, Chen YT, Dai SJ, Zhou XJ, Yang YX, Lou JS, Ji LT, Bao YT, Xuan L, Lin LN, Li CY. Berberine improves inflammatory responses of diabetes mellitus in zucker diabetic fatty rats and insulin-resistant HepG2 cells through the PPM1B pathway. J Immunol Res 2020; 2020: 2141508
  PubMedSearch in Google Scholar
• 39 Krümmel B, Plötz T, Jörns A, Lenzen S, Mehmeti I. The central role of glutathione peroxidase 4 in the regulation of ferroptosis and its implications for pro-inflammatory cytokine-mediated beta-cell death. Biochim Biophys Acta Mol Basis Dis 2021; 1867: 166114
  CrossrefPubMedSearch in Google Scholar
• 40 Lv X, Zhao Y, Yang X, Han H, Ge Y, Zhang M, Zhang H, Zhang M, Chen L. Berberine potentiates insulin secretion and prevents β-cell dysfunction through the miR-204/SIRT1 signaling pathway. Front Pharmacol 2021; 12: 720866
  CrossrefPubMedSearch in Google Scholar

• Supplementary Material