Peroxisome Proliferator Activated Receptor-gamma P465L Point Mutation in Diabetes and Atherosclerosis

The dominant-negative P467L mutation in peroxisome proliferator activated receptor-[gamma] (PPAR[gamma]) was identified in insulin resistant patients with hyperglycemia and lipodystrophy. However, mice carrying the corresponding Pparg-P465L mutation have normal insulin sensitivity, with mild hyperinsulinemia. I hypothesized that murine Pparg-P465L mutation leads to covert insulin resistance, masked by hyperinsulinemia, to retain normal plasma glucose. I introduced in PpargP465L/+ mice, Ins2-Akita mutation that causes islet apoptosis to lower plasma insulin. Unlike Ins2Akita/+ littermates, male PpargP465L/+Ins2Akita/+mice have drastically reduced life-span. PpargP465L/+Ins2Akita/+ females have aggravated hyperglycemia, smaller islets, reduced plasma insulin and impaired insulin sensitivity. Enhanced gluoconeogenesis, due to reduced plasma insulin, in PpargP465L/+Ins2Akita/+ females compared to Ins2Akita/+ littermates, contributes to their fasting hyperglycemia. While liver and skeletal muscles remain sensitive, adipose tissue is resistant to insulin. PpargP465L/+Ins2Akita/+ mice have smaller fat depots composed of larger adipocytes suggesting impaired lipid storage with subsequent hepatomegaly and hypertriglyceridemia. The increasing prevalence of obesity and insulin resistance and its negative impact on atherosclerotic cardiovascular disease is of major concern. In the second part of my study, I hypothesized that the L/+ mutation will worsen atherosclerosis in Apoe-/- mice. Despite a consistent increase in blood pressure, PPAR[gamma]P465L/+ mutation did not affect plaque size in Apoe-/- mice fed normal chow or high-fat-diet. Gene expression studies revealed significantly increased ABCA-1 and decreased CD-36 in PPAR[gamma]P465L/+;Apoe-/- macrophages compared to Apoe-/- littermates, suggesting reduced lipid accumulation. Cultured PPAR[gamma]P465L/+;Aoe-/- macrophages, had a significant reduction in DiI-labeled-VLDL uptake, compared to Apoe-/- macrophages. It is likely that balance between increased blood pressure with its pro-atherogenic potential and macrophage athero-protective phenotype results in an unchanged plaque. Bone marrow transfer experiments to investigate the role of PPAR[gamma]P465L/+ mutation specifically in the macrophage showed a trend towards decrease in plaque size in mice reconstituted with PPAR[gamma]P465L/+;Apoe-/- macrophages. Thus, I have established the causative link between PPAR[gamma]P465L mutation and insulin resistance phenotype, consolidating the crucial role of PPAR[gamma] in diabetes. PPAR[gamma]P465L mutation, however, does not change the atherosclerotic plaque size in Apoe-/- mice, and warrants further investigation

Apolipoprotein E knock-out and knock-in mice: atherosclerosis, metabolic syndrome, and beyond: Fig. 1.

Given the multiple differences between mice and men, it was once thought that mice could not be used to model atherosclerosis, principally a human disease. Apolipoprotein E-deficient (apoEKO) mice have convincingly changed this view, and the ability to model human-like plaques in these mice has provided scientists a platform to study multiple facets of atherogenesis and to explore potential therapeutic interventions. In addition to its well-established role in lipoprotein metabolism, recent observations of reduced adiposity and improved glucose homeostasis in apoEKO mice suggest that apoE may also play a key role in energy metabolism in peripheral organs, including adipose tissue. Finally, along with apoEKO mice, knockin mice expressing human apoE isoforms in place of endogenous mouse apoE have provided insights into how quantitative and qualitative genetic alterations interact with the environment in the pathogenesis of complex human diseases

Further evidence for the involvement of EFL1 in a Shwachman-Diamond-like syndrome and expansion of the phenotypic features.

Recent evidence has implicated EFL1 in a phenotype overlapping Shwachman-Diamond syndrome (SDS), with the functional interplay between EFL1 and the previously known causative gene SBDS accounting for the similarity in clinical features. Relatively little is known about the phenotypes associated with pathogenic variants in the EFL1 gene, but the initial indication was that phenotypes may be more severe, when compared with SDS. We report a pediatric patient who presented with a metaphyseal dysplasia and was found to have biallelic variants in EFL1 on reanalysis of trio whole-exome sequencing data. The variant had not been initially reported because of the research laboratory's focus on de novo variants. Subsequent phenotyping revealed variability in her manifestations. Although her metaphyseal abnormalities were more severe than in the original reported cohort with EFL1 variants, the bone marrow abnormalities were generally mild, and there was equivocal evidence for pancreatic insufficiency. Despite the limited number of reported patients, variants in EFL1 appear to cause a broader spectrum of symptoms that overlap with those seen in SDS. Our report adds to the evidence of EFL1 being associated with an SDS-like phenotype and provides information adding to our understanding of the phenotypic variability of this disorder. Our report also highlights the value of exome data reanalysis when a diagnosis is not initially apparent

Peroxisome proliferator activated receptor-gamma P465L mutation in diabetes and atherosclerosis

The dominant-negative P467L mutation in peroxisome proliferator activated receptor-γ (PPARγ) was identified in insulin resistant patients with hyperglycemia and lipodystrophy. However, mice carrying the corresponding Pparg-P465L mutation have normal insulin sensitivity, with mild hyperinsulinemia. I hypothesized that murine Pparg-P465L mutation leads to covert insulin resistance, masked by hyperinsulinemia, to retain normal plasma glucose. I introduced in PpargP465L/+ mice, Ins2Akita mutation that causes islet apoptosis to lower plasma insulin. Unlike Ins2Akita/+ littermates, male PpargP465L/+Ins2Akita/+ mice have drastically reduced life-span. PpargP465L/+Ins2 Akita/+ females have aggravated hyperglycemia, smaller islets, reduced plasma insulin and impaired insulin sensitivity. Enhanced gluoconeogenesis, due to reduced plasma insulin, in PpargP465L/+Ins2 Akita/+ females compared to Ins2Akita/+ littermates, contributes to their fasting hyperglycemia. While liver and skeletal muscles remain sensitive, adipose tissue is resistant to insulin. PpargP465L/+Ins2Akita/+ mice have smaller fat depots composed of larger adipocytes suggesting impaired lipid storage with subsequent hepatomegaly and hypertriglyceridemia. The increasing prevalence of obesity and insulin resistance and its negative impact on atherosclerotic cardiovascular disease is of major concern. In the second part of my study, I hypothesized that the L/+ mutation will worsen atherosclerosis in Apoe-/- mice. Despite a consistent increase in blood pressure, PPARγP465L/+ mutation did not affect plaque size in Apoe-/- mice fed normal chow or high-fat-diet. Gene expression studies revealed significantly increased ABCA-1 and decreased CD-36 in PPARγP465L/+;Apoe-/- macrophages compared to Apoe-/- littermates, suggesting reduced lipid accumulation. Cultured PPARγP465L/+;Aoe-/- macrophages, had a significant reduction in DiI-labeled-VLDL uptake, compared to Apoe-/- macrophages. It is likely that balance between increased blood pressure with its pro-atherogenic potential and macrophage athero-protective phenotype results in an unchanged plaque. Bone marrow transfer experiments to investigate the role of PPARγP465L/+ mutation specifically in the macrophage showed a trend towards decrease in plaque size in mice reconstituted with PPARγP465L/+; Apoe-/- macrophages. Thus, I have established the causative link between PPARγP465L mutation and insulin resistance phenotype, consolidating the crucial role of PPARγ in diabetes. PPARγP465L mutation, however, does not change the atherosclerotic plaque size in Apoe-/- mice, and warrants further investigation

Primary Invasive Squamous Cell Carcinoma of the Nipple

Squamous cell carcinoma is one of the most common cutaneous cancers; however, primary squamous cell carcinoma of the nipple is extremely rare. Among the few reported cases, the majority have occurred in older women with rare cases seen in younger women and male patients. Our patient presented with an exophytic mass of the right nipple while pregnant. A superficial biopsy was reviewed at an outside institution and then at our institution and diagnosed as squamous papilloma and then as hyperkeratosis of the nipple, respectively. The subsequent excisional biopsy revealed multiple nests of tumor cells extending into the dermis with associated chronic inflammatory infiltrate, and the lesion was diagnosed as a primary invasive squamous cell carcinoma of the nipple. Following that, a wide local excision of the excision site and sampling of the regional lymph nodes were negative for carcinoma. Due to the rarity of this diagnosis, it is not known whether prognosis and response to therapy differ from cutaneous squamous cell carcinoma at other sites. Therefore, risk stratification and therapy have been based on those for cutaneous squamous cell carcinoma

Apolipoprotein E knock-out and knock-in mice: atherosclerosis, metabolic syndrome, and beyond

Given the multiple differences between mice and men, it was once thought that mice could not be used to model atherosclerosis, principally a human disease. Apolipoprotein E-deficient (apoEKO) mice have convincingly changed this view, and the ability to model human-like plaques in these mice has provided scientists a platform to study multiple facets of atherogenesis and to explore potential therapeutic interventions. In addition to its well-established role in lipoprotein metabolism, recent observations of reduced adiposity and improved glucose homeostasis in apoEKO mice suggest that apoE may also play a key role in energy metabolism in peripheral organs, including adipose tissue. Finally, along with apoEKO mice, knockin mice expressing human apoE isoforms in place of endogenous mouse apoE have provided insights into how quantitative and qualitative genetic alterations interact with the environment in the pathogenesis of complex human diseases

A noncanonical function of cGAMP in inflammasome priming and activation

Recognition of pathogen-associated molecular patterns and danger-associated molecular patterns by host cells is an important step in innate immune activation. The DNA sensor cyclic guanosine monophosphate-adenosine monophosphate (cGAMP) synthase (cGAS) binds to DNA and produces cGAMP, which in turn binds to stimulator of interferon genes (STING) to activate IFN-I. Here we show that cGAMP has a noncanonical function in inflammasome activation in human and mouse cells. Inflammasome activation requires two signals, both of which are activated by cGAMP. cGAMP alone enhances expression of inflammasome components through IFN-I, providing the priming signal. Additionally, when combined with a priming signal, cGAMP activates the inflammasome through an AIM2, NLRP3, ASC, and caspase-1 dependent process. These two cGAMP-mediated functions, priming and activation, have differential requirements for STING. Temporally, cGAMP induction of IFN-I precedes inflammasome activation, which then occurs when IFN-I is waning. In mice, cGAS/cGAMP amplify both inflammasome and IFN-I to control murine cytomegalovirus. Thus, cGAMP activates the inflammasome in addition to IFN-I, and activation of both is needed to control infection by a DNA virus

Further evidence for the involvement of EFL1 in a Shwachman-Diamond-like syndrome and expansion of the phenotypic features.

Recent evidence has implicated EFL1 in a phenotype overlapping Shwachman-Diamond syndrome (SDS), with the functional interplay between EFL1 and the previously known causative gene SBDS accounting for the similarity in clinical features. Relatively little is known about the phenotypes associated with pathogenic variants in the EFL1 gene, but the initial indication was that phenotypes may be more severe, when compared with SDS. We report a pediatric patient who presented with a metaphyseal dysplasia and was found to have biallelic variants in EFL1 on reanalysis of trio whole-exome sequencing data. The variant had not been initially reported because of the research laboratory's focus on de novo variants. Subsequent phenotyping revealed variability in her manifestations. Although her metaphyseal abnormalities were more severe than in the original reported cohort with EFL1 variants, the bone marrow abnormalities were generally mild, and there was equivocal evidence for pancreatic insufficiency. Despite the limited number of reported patients, variants in EFL1 appear to cause a broader spectrum of symptoms that overlap with those seen in SDS. Our report adds to the evidence of EFL1 being associated with an SDS-like phenotype and provides information adding to our understanding of the phenotypic variability of this disorder. Our report also highlights the value of exome data reanalysis when a diagnosis is not initially apparent