| 1. |
- Azhar, Salman, et al.
(författare)
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Novel ABCA1 peptide agonists with antidiabetic action
- 2019
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Ingår i: Molecular and Cellular Endocrinology. - : Elsevier BV. - 0303-7207. ; 480, s. 1-11
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Tidskriftsartikel (refereegranskat)abstract
- Previously, apoE-derived ABCA1 agonist peptides have been shown to possess anti-atherosclerotic and possibly antidiabetic properties. Here we assessed the in vitro and in vivo actions of a second generation of ABCA1 peptide agonists, CS6253 and T6991-2, on glucose homeostasis. The results show that these two peptides improve glucose tolerance in a prediabetic diet-induced obesity mouse model by enhancing insulin secretion. It was further demonstrated that T6991-2 also improved glucose tolerance in leptin-deficient (ob/ob) mice. CS6253 increased insulin secretion both under basal conditions and in response to high glucose stimulation in pancreatic INS-1 β-cells rendered leptin receptor deficient with specific siRNA. Additional in vitro cell studies suggest that the CS6253 agonist attenuates hepatic gluconeogenesis and glucose transport. It also potentiates insulin-stimulated glucose uptake and utilization. These observed anti-diabetic actions suggest additional benefits of the CS6253 and T6991-2 ABCA1 peptide agonists for cardiovascular disease beyond their direct anti-atherosclerosis properties previously described.
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| 2. |
- Del Giudice, Rita, et al.
(författare)
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High-efficient bacterial production of human ApoA-I amyloidogenic variants
- 2018
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Ingår i: Protein Science. - : Wiley. - 0961-8368. ; 27:12, s. 2101-2109
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Tidskriftsartikel (refereegranskat)abstract
- Apolipoprotein A-I (ApoA-I)-related amyloidosis is a rare disease caused by missense mutations in the APOA1 gene. These mutations lead to protein aggregation and abnormal accumulation of ApoA-I amyloid fibrils in heart, liver, kidneys, skin, nerves, ovaries, or testes. Consequently, the carriers are at risk of single- or multi-organ failure and of need of organ transplantation. Understanding the basic molecular structure and function of ApoA-I amyloidogenic variants, as well as their biological effects, is, therefore, of great interest. However, the intrinsic low stability of this type of proteins makes their overexpression and purification difficult. To overcome this barrier, we here describe an optimized production and purification procedure for human ApoA-I amyloidogenic proteins that efficiently provides between 46 mg and 91 mg (depending on the protein variant) of pure protein per liter of Escherichia coli culture. Structural integrity of the amyloidogenic and native ApoA-I proteins were verified by circular dichroism spectroscopy and intrinsic fluorescence analysis, and preserved functionality was demonstrated by use of a lipid clearance assay as well as by reconstitution of high-density lipoprotein (HDL) particles. In conclusion, the use of the described high-yield protein production system to obtain amyloidogenic ApoA-I proteins, and their native counterpart, will enable molecular and cellular experimental studies aimed to explain the molecular basis for this rare disease.
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| 3. |
- Del Giudice, Rita, et al.
(författare)
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Structural determinants in ApoA-I amyloidogenic variants explain improved cholesterol metabolism despite low HDL levels
- 2017
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Ingår i: Biochimica et Biophysica Acta. - : Elsevier BV. - 0006-3002. ; 1863:12, s. 3038-3048
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Tidskriftsartikel (refereegranskat)abstract
- Twenty Apolipoprotein A-I (ApoA-I) variants are responsible for a systemic hereditary amyloidosis in which protein fibrils can accumulate in different organs, leading to their failure. Several ApoA-I amyloidogenic mutations are also associated with hypoalphalipoproteinemia, low ApoA-I and high-density lipoprotein (HDL)-cholesterol plasma levels; however, subjects affected by ApoA-I-related amyloidosis do not show a higher risk of cardiovascular diseases (CVD). The structural features, the lipid binding properties and the functionality of four ApoA-I amyloidogenic variants were therefore inspected in order to clarify the paradox observed in the clinical phenotype of the affected subjects. Our results show that ApoA-I amyloidogenic variants are characterized by a different oligomerization pattern and that the position of the mutation in the ApoA-I sequence affects the molecular structure of the formed HDL particles. Although lipidation increases ApoA-I proteins stability, all the amyloidogenic variants analyzed show a lower affinity for lipids, both in vitro and in ex vivo mouse serum. Interestingly, the lower efficiency at forming HDL particles is compensated by a higher efficiency at catalysing cholesterol efflux from macrophages. The decreased affinity of ApoA-I amyloidogenic variants for lipids, together with the increased efficiency in the cholesterol efflux process, could explain why, despite the unfavourable lipid profile, patients affected by ApoA-I related amyloidosis do not show a higher CVD risk.
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| 4. |
- Domingo-Espín, Joan, et al.
(författare)
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Dual actions of apolipoprotein A-I on glucose-stimulated insulin secretion and insulin-independent peripheral tissue glucose uptake lead to increased heart and skeletal muscle glucose disposal
- 2016
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Ingår i: Diabetes. - : American Diabetes Association. - 0012-1797 .- 1939-327X. ; 65:7, s. 1838-1848
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Tidskriftsartikel (refereegranskat)abstract
- Apolipoprotein A-I (apoA-I) of HDL is central to the transport of cholesterol in circulation. ApoA-I also provides glucose control with described in vitro effects of apoA-I on β-cell insulin secretion and muscle glucose uptake. In addition, apoA-I injections in insulin-resistant diet-induced obese (DIO) mice lead to increased glucose-stimulated insulin secretion (GSIS) and peripheral tissue glucose uptake. However, the relative contribution of apoA-I as an enhancer of GSIS in vivo and as a direct stimulator of insulin-independent glucose uptake is not known. Here, DIO mice with instant and transient blockade of insulin secretion were used in glucose tolerance tests and in positron emission tomography analyses. Data demonstrate that apoA-I to an equal extent enhances GSIS and acts as peripheral tissue activator of insulin-independent glucose uptake and verify skeletal muscle as an apoA-I target tissue. Intriguingly, our analyses also identify the heart as an important target tissue for the apoA-I-stimulated glucose uptake, with potential implications in diabetic cardiomyopathy. Explorations of apoA-I as a novel antidiabetic drug should extend to treatments of diabetic cardiomyopathy and other cardiovascular diseases in patients with diabetes.
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| 5. |
- Domingo-Espín, Joan, et al.
(författare)
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Site-specific glycations of apolipoprotein A-I lead to differentiated functional effects on lipid-binding and on glucose metabolism
- 2018
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Ingår i: Biochimica et Biophysica Acta - Molecular Basis of Disease. - : Elsevier BV. - 0925-4439. ; 1864:9, s. 2822-2834
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Tidskriftsartikel (refereegranskat)abstract
- Prolonged hyperglycemia in poorly controlled diabetes leads to an increase in reactive glucose metabolites that covalently modify proteins by non-enzymatic glycation reactions. Apolipoprotein A-I (apoA-I) of high-density lipoprotein (HDL) is one of the proteins that becomes glycated in hyperglycemia. The impact of glycation on apoA-I protein structure and function in lipid and glucose metabolism were investigated. ApoA-I was chemically glycated by two different glucose metabolites (methylglyoxal and glycolaldehyde). Synchrotron radiation and conventional circular dichroism spectroscopy were used to study apoA-I structure and stability. The ability to bind lipids was measured by lipid-clearance assay and native gel analysis, and cholesterol efflux was measured by using lipid-laden J774 macrophages. Diet induced obese mice with established insulin resistance, L6 rat and C2C12 mouse myocytes, as well as INS-1E rat insulinoma cells, were used to determine in vivo and in vitro glucose uptake and insulin secretion. Site-specific, covalent modifications of apoA-I (lysines or arginines) led to altered protein structure, reduced lipid binding capability and a reduced ability to catalyze cholesterol efflux from macrophages, partly in a modification-specific manner. The stimulatory effects of apoA-I on the in vivo glucose clearance were negatively affected when apoA-I was modified with methylglyoxal, but not with glycolaldehyde. The in vitro data showed that both glucose uptake in muscle cells and insulin secretion from beta cells were affected. Taken together, glycation modifications impair the apoA-I protein functionality in lipid and glucose metabolism, which is expected to have implications for diabetes patients with poorly controlled blood glucose.
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| 6. |
- Dwivedi, Om Prakash, et al.
(författare)
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Loss of ZnT8 function protects against diabetes by enhanced insulin secretion
- 2019
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Ingår i: Nature Genetics. - : Springer Science and Business Media LLC. - 1061-4036 .- 1546-1718. ; , s. 1-22
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Tidskriftsartikel (refereegranskat)abstract
- A rare loss-of-function allele p.Arg138* in SLC30A8 encoding the zinc transporter 8 (ZnT8), which is enriched in Western Finland, protects against type 2 diabetes (T2D). We recruited relatives of the identified carriers and showed that protection was associated with better insulin secretion due to enhanced glucose responsiveness and proinsulin conversion, particularly when compared with individuals matched for the genotype of a common T2D-risk allele in SLC30A8, p.Arg325. In genome-edited human induced pluripotent stem cell (iPSC)-derived β-like cells, we establish that the p.Arg138* allele results in reduced SLC30A8 expression due to haploinsufficiency. In human β cells, loss of SLC30A8 leads to increased glucose responsiveness and reduced KATP channel function similar to isolated islets from carriers of the T2D-protective allele p.Trp325. These data position ZnT8 as an appealing target for treatment aimed at maintaining insulin secretion capacity in T2D.
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| 7. |
- Edmunds, Shelley J, et al.
(författare)
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ApoAI-derived peptide increases glucose tolerance and prevents formation of atherosclerosis in mice
- 2019
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Ingår i: Diabetologia. - : Springer Science and Business Media LLC. - 1432-0428 .- 0012-186X. ; 62:7, s. 1257-1267
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Tidskriftsartikel (refereegranskat)abstract
- AIMS/HYPOTHESIS: Finding new treatment alternatives for individuals with diabetes with severe insulin resistance is highly desired. To identify novel mechanisms that improve glucose uptake in skeletal muscle, independently from insulin levels and signalling, we have explored the therapeutic potential of a short peptide sequence, RG54, derived from apolipoprotein A-I (ApoA-I).METHODS: INS-1E rat clonal beta cells, C2C12 rat muscle myotubes and J774 mouse macrophages were used to study the impact of RG54 peptide on glucose-stimulated insulin secretion, glucose uptake and cholesterol efflux, respectively. GTTs were carried out on diet-induced insulin-resistant and Leprdb diabetic mouse models treated with RG54 peptide, and the impact of RG54 peptide on atherosclerosis was evaluated in Apoe-/- mice. Control mice received ApoA-I protein, liraglutide or NaCl.RESULTS: The synthetic RG54 peptide induced glucose uptake in cultured muscle myotubes by a similar amount as insulin, and also primed pancreatic beta cells for improved glucose-stimulated insulin secretion. The findings were verified in diet-induced insulin-resistant and Leprdb diabetic mice, jointly confirming the physiological effect. The RG54 peptide also efficiently catalysed cholesterol efflux from macrophages and prevented the formation of atherosclerotic plaques in Apoe-/- mice.CONCLUSIONS/INTERPRETATION: The RG54 peptide exhibits good prospects for providing glucose control and reducing the risk of cardiovascular disease in individuals with severe insulin resistance.
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| 8. |
- Giudice, Rita Del, et al.
(författare)
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Synchrotron radiation circular dichroism spectroscopy reveals structural divergences in HDL-bound apoA-I variants
- 2017
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Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 7:1
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Tidskriftsartikel (refereegranskat)abstract
- Apolipoprotein A-I (apoA-I) in high-density lipoprotein (HDL) provides cardiovascular protection. Synchrotron radiation circular dichroism (SRCD) spectroscopy was used to analyze the dynamic solution structure of the apoA-I protein in the apo- and HDL-states and the protein structure conversion in HDL formation. Wild-type apoA-I protein was compared to human variants that either are protective (R173C, Milano) or lead to increased risk for ischaemic heart disease (A164S). Comparable secondary structure distributions in the HDL particles, including significant levels of beta strand/turn, were observed. ApoA-I Milano in HDL displayed larger size heterogeneity, increased protein flexibility, and an altered lipid-binding profile, whereas the apoA-I A164S in HDL showed decrease thermal stability, potentially linking the intrinsic HDL propensities of the variants to disease risk.
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| 9. |
- Petrlova, Jitka, et al.
(författare)
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Molecular crowding impacts the structure of apolipoprotein A-I with potential implications on in vivo metabolism and function
- 2016
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Ingår i: Biopolymers. - : Wiley. - 0006-3525. ; 105:10, s. 683-692
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Tidskriftsartikel (refereegranskat)abstract
- The effect molecular crowding, defined as the volume exclusion exerted by one soluble inert molecule upon another soluble molecule, has on the structure and self-interaction of lipid-free apoA-I were explored. The influence of molecular crowding on lipid-free apoA-I oligomerization and internal dynamics has been analyzed using electron paramagnetic resonance (EPR) spectroscopy measurements of nitroxide spin label at selected positions throughout the protein sequence and at varying concentrations of the crowding agent Ficoll-70. The targeted positions include sites previously shown to be sensitive for detecting intermolecular interaction via spin–spin coupling. Circular dichroism was used to study secondary structural changes in lipid-free apoA-I imposed by increasing concentrations of the crowding agent. Crosslinking and SDS-PAGE gel analysis was employed to further characterize the role molecular crowding plays in inducing apoA-I oligomerization. It was concluded that the dynamic apoA-I structure and oligomeric state was altered in the presence of the crowding agent. It was also found that the C-terminal was slightly more sensitive to molecular crowding. Finally, the data described the region around residue 217 in the C-terminal domain of apoA-I as the most sensitive reporter of the crowding-induced self-association of apoA-I. The implications of this behavior to in vivo functionality are discussed.
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| 10. |
- Svensson, Daniel, et al.
(författare)
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Apolipoprotein A-I attenuates LL-37-induced endothelial cell cytotoxicity
- 2017
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Ingår i: Biochemical and Biophysical Research Communications. - : Elsevier BV. - 0006-291X. ; 493:1, s. 71-76
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Tidskriftsartikel (refereegranskat)abstract
- The human cathelicidin peptide LL-37 has antimicrobial and anti-biofilm functions, but LL-37 may also damage the host by triggering inflammation and exerting a cytotoxic effect, thereby reducing host cell viability. Human plasma mitigates LL-37-induced host cell cytotoxicity but the underlying mechanisms are not completely understood. Apolipoprotein A-I (ApoA-I) is a plasma protein endowed with atheroprotective effects. Here, we investigate the interaction between ApoA-I and LL-37 by biochemical techniques, and furthermore assess if ApoA-I protects against LL-37-evoked cytotoxicity in human umbilical vein endothelial cells (HUVEC). Our results demonstrated that ApoA-I effectively binds LL-37. The binding of ApoA-I to LL-37 resulted in a structural rearrangement of the protein, but this interaction did not cause lower ApoA-I stability. Recombinant ApoA-I protected against LL-37-induced cytotoxicity in HUVEC and endogenous ApoA-I knockdown in HepG2 cells made the cells more sensitive to LL-37-evoked cytotoxicity. We conclude that ApoA-I physically interacts with LL-37 and antagonizes LL-37-induced down-regulation of endothelial cell viability suggesting that this mechanism counteracts endothelial cell dysfunction.
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