De novo construction of T cell compartment in humanized mice engrafted with iPSC-derived thymus organoids.

Hematopoietic humanized (hu) mice are powerful tools for modeling the action of human immune system and are widely used for preclinical studies and drug discovery. However, generating a functional human T cell compartment in hu mice remains challenging, primarily due to the species-related differences between human and mouse thymus. While engrafting human fetal thymic tissues can support robust T cell development in hu mice, tissue scarcity and ethical concerns limit their wide use. Here, we describe the tissue engineering of human thymus organoids from inducible pluripotent stem cells (iPSC-thymus) that can support the de novo generation of a diverse population of functional human T cells. T cells of iPSC-thymus-engrafted hu mice could mediate both cellular and humoral immune responses, including mounting robust proinflammatory responses on T cell receptor engagement, inhibiting allogeneic tumor graft growth and facilitating efficient Ig class switching. Our findings indicate that hu mice engrafted with iPSC-thymus can serve as a new animal model to study human T cell-mediated immunity and accelerate the translation of findings from animal studies into the clinic.

Mesenteric lymph nodes as alternative site for pancreatic islet transplantation in a diabetic rat model.

BACKGROUND: Islet transplantation has progressively become a safe alternative to pancreas transplantation for the treatment of type 1 diabetes. However, the long-term results of islet transplantation could be significantly increased by improving the quality of the islet isolation technique even exploring alternative islet transplantation sites to reduce the number of islets required to mitigate hyperglycemia. The goal of the study was to test the lymph node as a suitable anatomical location for islet engraftment in a rodent model. METHODS: Forty Lewis rats, 6-8 weeks old, body weight 250-300 g, have been used as islet donors and recipients in syngeneic islet transplantation experiments. Ten rats were rendered diabetic by one injection of 65 mg/Kg of streptozotocin. After pancreas retrieval from non diabetic donors, islet were isolated and transplanted in the mesenteric lymph nodes of 7 diabetic rats. Rats were followed for 30 days after islet transplantation. RESULTS: A total of 7 islet transplantations in mesenteric lymph nodes have been performed. Two rats died 24 and 36 h after transplantation due to complications. No transplanted rat acquired normal glucose blood levels and insulin independence after the transplantation. However, the mean blood levels of glycemia were significantly lower in transplanted rats compared with diabetic rats (470.4 mg/dl vs 605 mg/dl, p 0.04). Interestingly, transplanted rats have a significant weight increase after transplantation compared to diabetic rats (mean value 295 g in transplanted rats vs 245 g in diabetic rats, p < 0.05), with an overall improvement of social activities and health. Immunohistochemical analysis of the 5 mesenteric lymph nodes of transplanted rats demonstrated the presence of living islets in one lymph node. CONCLUSIONS: Although islet engraftment in lymph nodes is possible, islet transplantation in lymph nodes in rats resulted in few improvements of glucose parameters.

Inherent ER stress in pancreatic islet ß cells causes self-recognition by autoreactive T cells in type 1 diabetes.

Type 1 diabetes (T1D) is an autoimmune disease characterized by pancreatic ß cell destruction induced by islet reactive T cells that have escaped central tolerance. Many physiological and environmental triggers associated with T1D result in ß cell endoplasmic reticulum (ER) stress and dysfunction, increasing the potential for abnormal post-translational modification (PTM) of proteins. We hypothesized that ß cell ER stress induced by environmental and physiological conditions generates abnormally-modified proteins for the T1D autoimmune response. To test this hypothesis we exposed the murine CD4(+) diabetogenic BDC2.5 T cell clone to murine islets in which ER stress had been induced chemically (Thapsigargin). The BDC2.5 T cell IFNγ response to these cells was significantly increased compared to non-treated islets. This ß cell ER stress increased activity of the calcium (Ca(2+))-dependent PTM enzyme tissue transglutaminase 2 (Tgase2), which was necessary for full stress-dependent immunogenicity. Indeed, BDC2.5 T cells responded more strongly to their antigen after its modification by Tgase2. Finally, exposure of non-antigenic murine insulinomas to chemical ER stress in vitro or physiological ER stress in vivo caused increased ER stress and Tgase2 activity, culminating in higher BDC2.5 responses. Thus, ß cell ER stress induced by chemical and physiological triggers leads to ß cell immunogenicity through Ca(2+)-dependent PTM. These findings elucidate a mechanism of how ß cell proteins are modified and become immunogenic, and reveal a novel opportunity for preventing ß cell recognition by autoreactive T cells.

Bioengineering Thymus Organoids to Restore Thymic Function and Induce Donor-Specific Immune Tolerance to Allografts.

One of the major obstacles in organ transplantation is to establish immune tolerance of allografts. Although immunosuppressive drugs can prevent graft rejection to a certain degree, their efficacies are limited, transient, and associated with severe side effects. Induction of thymic central tolerance to allografts remains challenging, largely because of the difficulty of maintaining donor thymic epithelial cells in vitro to allow successful bioengineering. Here, the authors show that three-dimensional scaffolds generated from decellularized mouse thymus can support thymic epithelial cell survival in culture and maintain their unique molecular properties. When transplanted into athymic nude mice, the bioengineered thymus organoids effectively promoted homing of lymphocyte progenitors and supported thymopoiesis. Nude mice transplanted with thymus organoids promptly rejected skin allografts and were able to mount antigen-specific humoral responses against ovalbumin on immunization. Notably, tolerance to skin allografts was achieved by transplanting thymus organoids constructed with either thymic epithelial cells coexpressing both syngeneic and allogenic major histocompatibility complexes, or mixtures of donor and recipient thymic epithelial cells. Our results demonstrate the technical feasibility of restoring thymic function with bioengineered thymus organoids and highlight the clinical implications of this thymus reconstruction technique in organ transplantation and regenerative medicine.

Bioengineering mini functional thymic units with EAK16-II/EAKIIH6 self-assembling hydrogel.

Herein, we highlight the technical feasibility of generating a functional mini thymus with a novel hydrogel system, based on a peptide-based self-assembly platform that can induce the formation of 3-D thymic epithelial cell (TEC) clusters. Amphiphilic peptide EAK16-II co-assembled with its histidinylated analogue EAKIIH6 into beta-sheet fibrils. When adaptor complexes (recombinant protein A/G molecules loaded with both anti-His and anti-EpCAM IgGs) were added to the mix, TECs were tethered to the hydrogel and formed 3-D mini clusters. TECs bound to the hydrogel composites retained their molecular properties; and when transplanted into athymic nude mice, they supported the development of functional T-cells. These mini thymic units of TECs can be useful in clinical applications to reconstitute T-cell adaptive immunity.

In vitro exposure of pig neonatal isletlike cell clusters to human blood.

BACKGROUND: Pig islet grafts have been successful in treating diabetes in animal models. One remaining question is whether neonatal pig isletlike cell clusters (NICC) are resistant to the early loss of islets from the instant blood-mediated inflammatory reaction (IBMIR). METHODS: Neonatal isletlike cell clusters were harvested from three groups of piglets-(i) wild-type (genetically unmodified), (ii) α1,3-galactosyltransferase gene-knockout (GTKO)/CD46, and (iii) GTKO/CD46/CD39. NICC samples were mixed with human blood in vitro, and the following measurements were made-antibody binding; complement activation; speed of islet-induced coagulation; C-peptide; glutamic acid decarboxylase (GAD65) release; viability. RESULTS: Time to coagulation and viability were both reduced in all groups compared to freshly drawn non-anticoagulated human blood and autologous combinations, respectively. Antibody binding to the NICC occurred in all groups. CONCLUSIONS: Neonatal isletlike cell clusters were subject to humoral injury with no difference associated to their genetic characteristics.

Compromised central tolerance of ICA69 induces multiple organ autoimmunity.

For reasons not fully understood, patients with an organ-specific autoimmune disease have increased risks of developing autoimmune responses against other organs/tissues. We identified ICA69, a known ß-cell autoantigen in Type 1 diabetes, as a potential common target in multi-organ autoimmunity. NOD mice immunized with ICA69 polypeptides exhibited exacerbated inflammation not only in the islets, but also in the salivary glands. To further investigate ICA69 autoimmunity, two genetically modified mouse lines were generated to modulate thymic ICA69 expression: the heterozygous ICA69(del/wt) line and the thymic medullary epithelial cell-specific deletion Aire-ΔICA69 line. Suboptimal central negative selection of ICA69-reactive T-cells was observed in both lines. Aire-ΔICA69 mice spontaneously developed coincident autoimmune responses to the pancreas, the salivary glands, the thyroid, and the stomach. Our findings establish a direct link between compromised thymic ICA69 expression and autoimmunity against multiple ICA69-expressing organs, and identify a potential novel mechanism for the development of multi-organ autoimmune diseases.

Repopulation of decellularized organ scaffolds with human pluripotent stem cell-derived pancreatic progenitor cells.

Diabetes is an emerging global epidemic that affects more that 285 million people worldwide. Engineering of endocrine pancreas tissue holds great promise for the future of diabetes therapy. Here we demonstrate the feasibility of re-engineering decellularized organ scaffolds using regenerative cell source. We differentiated human pluripotent stem cells (hPSC) toward pancreatic progenitor (PP) lineage and repopulated decellularized organ scaffolds with these hPSC-PP cells. We observed that hPSCs cultured and differentiated as aggregates are more suitable for organ repopulation than isolated single cell suspension. However, recellularization with hPSC-PP aggregates require a more extensive vascular support, which was found to be superior in decellularized liver over the decellularized pancreas scaffolds. Upon continued culture for nine days with chemical induction in the bioreactor, the seeded hPSC-PP aggregates demonstrated extensive and uniform cellular repopulation and viability throughout the thickness of the liver scaffolds. Furthermore, the decellularized liver scaffolds was supportive of the endocrine cell fate of the engrafted cells. Our novel strategy to engineer endocrine pancreas construct is expected to find potential applications in preclinical testing, drug discovery and diabetes therapy.

Autologous Islet Transplantation After Total Pancreatectomy in a Patient Recovered from SARS-CoV-2: A Case Report.

BACKGROUND SARS-CoV-2 infection or COVID-19 disease has been linked to the onset of diabetes and metabolic dysregulation because it has been suggested that viral entry proteins, specifically ACE2 and TMPRSS2, are expressed in the exocrine cells and ductal epithelium of the pancreas. Because of the unknown effect this can have on islet function, there can be doubt that patients with previous SARS-CoV-2 infections are good candidates for autologous islet transplantation after total pancreatectomy (TPAIT). CASE REPORT A patient with a history of chronic pancreatitis and previous non-surgical interventions was presented as a viable candidate for TPAIT at our institution. Approximately 1 month later, the patient contracted a SARS-CoV-2 infection, resulting in a mild case of COVID-19. The infection resolved without the need for hospitalization. At the time of this occurrence, COVID-19 was primarily considered a respiratory ailment, and little was known of the potential association between metabolic dysfunction and SARS-CoV-2. Islet isolation and surgery proceeded in a textbook manner with no surgical complications. The patient was weaned off exogenous insulin within 3 months after transplantation. CONCLUSIONS Favorable outcomes after surgery included pain reduction, islet function, and improved quality of life for the patient in the first 6 months after the procedure. These successful results demonstrate that SARS-CoV-2 infection did not prevent the patient from achieving good glucose regulation after auto-islet transplantation. This outcome suggests that, at least in this instance of mild infection, there were no long-lasting negative COVID-19-associated effects on the transplanted islets that might impact islet function.

Dual pancreatic adrenergic and dopaminergic signaling as a therapeutic target of bromocriptine.

Bromocriptine is approved as a diabetes therapy, yet its therapeutic mechanisms remain unclear. Though bromocriptine's actions have been mainly attributed to the stimulation of brain dopamine D2 receptors (D2R), bromocriptine also targets the pancreas. Here, we employ bromocriptine as a tool to elucidate the roles of catecholamine signaling in regulating pancreatic hormone secretion. In ß-cells, bromocriptine acts on D2R and α2A-adrenergic receptor (α2A-AR) to reduce glucose-stimulated insulin secretion (GSIS). Moreover, in α-cells, bromocriptine acts via D2R to reduce glucagon secretion. α2A-AR activation by bromocriptine recruits an ensemble of G proteins with no ß-arrestin2 recruitment. In contrast, D2R recruits G proteins and ß-arrestin2 upon bromocriptine stimulation, demonstrating receptor-specific signaling. Docking studies reveal distinct bromocriptine binding to α2A-AR versus D2R, providing a structural basis for bromocriptine's dual actions on ß-cell α2A-AR and D2R. Together, joint dopaminergic and adrenergic receptor actions on α-cell and ß-cell hormone release provide a new therapeutic mechanism to improve dysglycemia.

Global deficits in development, function, and gene expression in the endocrine pancreas in a deletion mouse model of Prader-Willi syndrome.

Prader-Willi syndrome (PWS) is a multisystem disorder caused by genetic loss of function of a cluster of imprinted, paternally expressed genes. Neonatal failure to thrive in PWS is followed by childhood-onset hyperphagia and obesity among other endocrine and behavioral abnormalities. PWS is typically assumed to be caused by an unknown hypothalamic-pituitary dysfunction, but the underlying pathogenesis remains unknown. A transgenic deletion mouse model (TgPWS) has severe failure to thrive, with very low levels of plasma insulin and glucagon in fetal and neonatal life prior to and following onset of progressive hypoglycemia. In this study, we tested the hypothesis that primary deficits in pancreatic islet development or function may play a fundamental role in the TgPWS neonatal phenotype. Major pancreatic islet hormones (insulin, glucagon) were decreased in TgPWS mice, consistent with plasma levels. Immunohistochemical analysis of the pancreas demonstrated disrupted morphology of TgPWS islets, with reduced α- and ß-cell mass arising from an increase in apoptosis. Furthermore, in vivo and in vitro studies show that the rate of insulin secretion is significantly impaired in TgPWS ß-cells. In TgPWS pancreas, mRNA levels for genes encoding all pancreatic hormones, other secretory factors, and the ISL1 transcription factor are upregulated by either a compensatory response to plasma hormone deficiencies or a primary effect of a deleted gene. Our findings identify a cluster of imprinted genes required for the development, survival, coordinate regulation of genes encoding hormones, and secretory function of pancreatic endocrine cells, which may underlie the neonatal phenotype of the TgPWS mouse model.

Pig-to-nonhuman primates pancreatic islet xenotransplantation: an overview.

The therapy of type 1 diabetes is an open challenging problem. The restoration of normoglycemia and insulin independence in immunosuppressed type 1 diabetic recipients of islet allotransplantation has shown the potential of a cell-based diabetes therapy. Even if successful, this approach poses a problem of scarce tissue supply. Xenotransplantation can be the answer to this limited donor availability and, among possible candidate tissues for xenotransplantation, porcine islets are the closest to a future clinical application. Xenotransplantation, with pigs as donors, offers the possibility of using healthy, living, and genetically modified islets from pathogen-free animals available in unlimited number of islets. Several studies in the pig-to-nonhuman primate model demonstrated the feasibility of successful preclinical islet xenotransplantation and have provided insights into the critical events and possible mechanisms of immune recognition and rejection of xenogeneic islet grafts. Particularly promising results in the achievement of prolonged insulin independence were obtained with newly developed, genetically modified pigs islets able to produce immunoregulatory products, using different implantation sites, and new immunotherapeutic strategies. Nonetheless, further efforts are needed to generate additional safety and efficacy data in nonhuman primate models to safely translate these findings into the clinic.

Suppression of autoimmune diabetes by soluble galectin-1.

Type 1 diabetes (T1D) is a T cell-mediated autoimmune disease that targets the beta-cells of the pancreas. We investigated the ability of soluble galectin-1 (gal-1), an endogenous lectin that promotes T cell apoptosis, to down-regulate the T cell response that destroys the pancreatic beta-cells. We demonstrated that in nonobese diabetic (NOD) mice, gal-1 therapy reduces significantly the amount of Th1 cells, augments the number of T cells secreting IL-4 or IL-10 specific for islet cell Ag, and causes peripheral deletion of beta-cell-reactive T cells. Administration of gal-1 prevented the onset of hyperglycemia in NOD mice at early and subclinical stages of T1D. Preventive gal-1 therapy shifted the composition of the insulitis into an infiltrate that did not invade the islets and that contained a significantly reduced number of Th1 cells and a higher percentage of CD4(+) T cells with content of IL-4, IL-5, or IL-10. The beneficial effects of gal-1 correlated with the ability of the lectin to trigger apoptosis of the T cell subsets that cause beta-cell damage while sparing naive T cells, Th2 lymphocytes, and regulatory T cells in NOD mice. Importantly, gal-1 reversed beta-cell autoimmunity and hyperglycemia in NOD mice with ongoing T1D. Because gal-1 therapy did not cause major side effects or beta-cell toxicity in NOD mice, the use of gal-1 to control beta-cell autoimmunity represents a novel alternative for treatment of subclinical or ongoing T1D.

Human Hemangioblast-Derived Mesenchymal Stem Cells Promote Islet Engraftment in a Minimal Islet Mass Transplantation Model in Mice.

Islet transplantation can restore glycemic control in patients with type 1 diabetes. Using this procedure, the early stages of engraftment are often crucial to long-term islet function, and outcomes are not always successful. Numerous studies have shown that mesenchymal stem cells (MSCs) facilitate islet graft function. However, experimental data can be inconsistent due to variables associated with MSC generation (including donor characteristics and tissue source), thus, demonstrating the need for a well-characterized and uniform cell product before translation to the clinic. Unlike bone marrow- or adipose tissue-derived MSCs, human embryonic stem cell-derived-MSCs (hESC-MSCs) offer an unlimited source of stable and highly-characterized cells that are easily scalable. Here, we studied the effects of human hemangioblast-derived mesenchymal cells (HMCs), (i.e., MSCs differentiated from hESCs using a hemangioblast intermediate), on islet cell transplantation using a minimal islet mass model. The co-transplantation of the HMCs allowed a mass of islets that was insufficient to correct diabetes on its own to restore glycemic control in all recipients. Our in vitro studies help to elucidate the mechanisms including reduction of cytokine stress by which the HMCs support islet graft protection in vivo. Derivation, stability, and scalability of the HMC source may offer unique advantages for clinical applications, including fewer islets needed for successful islet transplantation.

Dopamine regulates pancreatic glucagon and insulin secretion via adrenergic and dopaminergic receptors.

Dopamine (DA) and norepinephrine (NE) are catecholamines primarily studied in the central nervous system that also act in the pancreas as peripheral regulators of metabolism. Pancreatic catecholamine signaling has also been increasingly implicated as a mechanism responsible for the metabolic disturbances produced by antipsychotic drugs (APDs). Critically, however, the mechanisms by which catecholamines modulate pancreatic hormone release are not completely understood. We show that human and mouse pancreatic α- and ß-cells express the catecholamine biosynthetic and signaling machinery, and that α-cells synthesize DA de novo. This locally-produced pancreatic DA signals via both α- and ß-cell adrenergic and dopaminergic receptors with different affinities to regulate glucagon and insulin release. Significantly, we show DA functions as a biased agonist at α2A-adrenergic receptors, preferentially signaling via the canonical G protein-mediated pathway. Our findings highlight the interplay between DA and NE signaling as a novel form of regulation to modulate pancreatic hormone release. Lastly, pharmacological blockade of DA D2-like receptors in human islets with APDs significantly raises insulin and glucagon release. This offers a new mechanism where APDs act directly on islet α- and ß-cell targets to produce metabolic disturbances.

Organ-specific ECM arrays for investigating cell-ECM interactions during stem cell differentiation.

Pluripotent stem cells are promising source of cells for tissue engineering, regenerative medicine and drug discovery applications. The process of stem cell differentiation is regulated by multi-parametric cues from the surrounding microenvironment, one of the critical one being cell interaction with extracellular matrix (ECM). The ECM is a complex tissue-specific structure which is an important physiological regulator of stem cell function and fate. Recapitulating this native ECM microenvironment niche is best facilitated by decellularized tissue/organ derived ECM, which can faithfully reproduce the physiological environment with high fidelity toin vivocondition and promote tissue-specific cellular development and maturation. Recognizing the need for organ specific ECM in a 3D culture environment in driving phenotypic differentiation and maturation of hPSCs, we fabricated an ECM array platform using native-mimicry ECM from decellularized organs (namely pancreas, liver and heart), which allows cell-ECM interactions in both 2D and 3D configuration. The ECM array was integrated with rapid quantitative imaging for a systematic investigation of matrix protein profiles and sensitive measurement of cell-ECM interaction during hPSC differentiation. We tested our platform by elucidating the role of the three different organ-specific ECM in supporting induced pancreatic differentiation of hPSCs. While the focus of this report is on pancreatic differentiation, the developed platform is versatile to be applied to characterize any lineage specific differentiation.

Pig-to-Macaque Islet Xenotransplantation.

The advancement toward a clinical application for porcine islets to cure diabetes in humans must include reproducible long-term successes in non-human primate (NHP) models. Many dedicated researchers around the world are continuing to work toward this goal. In this chapter, we describe procedures for islet isolation of pancreatic islets from adult and neonatal/fetal pigs. We further include procedures for the induction of diabetes in non-human primates and subsequent insulin therapy, islet transplantation, immunosuppression, and also the daily maintenance of xenotransplanted NHPs. The procedures that we outline in this chapter are ones that we have successfully utilized in pig-to-NHP islet transplantation models. However, where appropriate, alternative methods will also be identified.

A Method for Islet Transplantation to the Omentum in Mouse.

Islet transplantation has been proposed to be a potential treatment for type 1 diabetes. Recent compelling evidence indicates that intravascular islet infusion is far from ideal and therefore, the omentum is re-emerging as a potentially valuable site for islet transplantation. This experiment requires the isolation of high quality islets and the implantation of the islets to the diabetic recipients. Transplantation to the omentum requires surgical steps that can be better demonstrated visually. Here, the detailed steps for this procedure are presented. Two methods of mixing the isolated islets with hydrogel before placing the mixture into the omental pouch of diabetic mice are described here. Different hydrogels are used for the different conditions. Blood glucose levels of diabetic mouse recipients of syngeneic islets in the omentum were monitored for up to 35 days. Some animals were sacrificed after 14 days to perform immuno-histochemical analysis. This pre-clinical transplantation approach can be used as preliminary data leading up to translation to clinical transplantation.

The Future of Islet Transplantation Is Now.

Milestones in the history of diabetes therapy include the discovery of insulin and successful methods of beta cell replacement including whole pancreas and islet cell transplantation options. While pancreas transplantation remains the gold standard for patients who have difficulty controlling their symptoms with exogenous insulin, islet allotransplantation is now able to provide similar results with some advantages that make it an attractive potential alternative. The Edmonton Protocol, which incorporated a large dose of islets from multiple donors with steroid-free immunosuppression helped to establish the modern era of islet transplantation almost 20 years ago. While islet allotransplantation is recognized around the world as a powerful clinical therapy for type 1 diabetes it is not yet recognized by the Federal Drug Administration of the United States. Large-scale clinical trials administered by the Clinical Islet Transplantation Consortium have recently demonstrated that the well-regulated manufacture of a human islet product transplanted into patients with difficult to control type 1 diabetes and with a history of severe hyperglycemic episodes can safely and efficaciously maintain glycemic balance and eliminate the most severe complications associated with diabetes. The results of these clinical trials have established a strong basis for licensure of clinical islet allotransplantation in the US. Recognition by the Federal Drug Administration would likely lead to third party reimbursement for islet allotransplantation as a therapeutic option in the United States and would make the treatment available to many more patients. The high costs of rampant diabetes justify the expense of the treatment, which is in-line with the costs of clinical pancreas transplantation. While much enthusiasm and hope is raised toward the development and optimization of stem cell therapy, the islet transplantation community should push toward licensure, if that means broader access of this procedure to patients who may benefit from it. Even as we prepare to take the first steps in that direction, we must acknowledge the new challenges that a shift from the experimental to clinical will bring. Clinical islet allotransplantation in the United States would be a game-changing event in the treatment of type 1 diabetes and also generate enthusiasm for continued research.

Lipin1 regulation by estrogen in uterus and liver: implications for diabetes and fertility.

Estrogens are essential for fertility and also have important effects on regulation of adiposity and the euglycemic state. We report here that lipin1, a candidate gene for lipodystrophy and obesity that is a phosphatidic acid phosphatase critical in regulation of cellular levels of diacylglycerol and triacylglycerol and a key regulator of lipid utilization, is rapidly and robustly down-regulated in the uterus by estradiol via the estrogen receptor. Lipin1 is expressed predominantly in the uterine luminal and glandular epithelium, and during the estrous cycle, lipin1 is lowest when blood levels of estrogen are highest. Lipin1 is expressed throughout all cells in the liver of ovariectomized female mice, and a sustained down-regulation is observed at the mRNA, protein and immunohistochemical levels after estrogen administration. Because the coupling of proper energy use and availability is central to reproduction, we also investigated expression of lipin1 in the uterus and liver of several mouse models of diabetes. Nonobese diabetic (NOD) mice, which have high blood levels of estrogen and impaired fertility, were severely deficient in lipin1 in the uterus and liver, which, interestingly, could be restored by insulin treatment. By contrast, nonobese diabetic/severe combined immunodeficient (NOD-SCID) mice, which do not develop diabetes, showed normal levels of lipin1. Our findings of lipin1 regulation by estrogen in two key target organs suggest a new role for this lipid-regulating phosphatase not only in central metabolic regulation but also in uterine function and reproductive biology. Estrogen regulation of lipin1 may provide a mechanistic link between estrogens, lipid metabolism, and lipid signaling.