Synthesis of N-Hydroxysuccinimide Esters, N-Acylsaccharins, and Activated Esters from Carboxylic Acids Using I2/PPh3.

A method for the syntheses of isolable, active esters is described in which carboxylic acids are treated with triphenylphosphine, iodine, and triethylamine. Active esters accessible in this way include N-hydroxysuccinimide esters, N-hydroxyphthalimide esters (N-(acyloxy)phthalimides), N-acylsaccharins, pentafluorophenol esters, pentachlorophenol esters, N-hydroxybenzotriazole esters, and hexafluoro-2-propanol esters. The approach can be similarly applied toward the formation of N-acylsaccharins and N-acylimidazoles. The method is suitable for the formation of isolable active esters of aromatic and aliphatic activated acids as well as α-amino acid derivatives. These products are widely used reagents in organic synthesis, peptide synthesis, medicinal chemistry, and chemical biology (e.g., for bioconjugations). The method has broad substrate scope, uses simple and inexpensive reagents, avoids the use of carbodiimides or other coupling agents, and occurs at room temperature. Additionally, the diastereomers of compound Boc-Ala-NHCHPh are demonstrated to be distinguishable by 1H NMR (in DMSO-d6), allowing for a straightforward NMR method to establish the degree of racemization of activated esters of Boc-Ala or amide bond formations using Boc-Ala.

Total Synthesis of the 2,5-Disubstituted γ-Pyrone E1 UAE Inhibitor Himeic Acid A.

The first total synthesis of the E1 ubiquitin-activating enzyme inhibitor, himeic acid A, is reported. A McCombie reaction was used to form the core γ-pyrone via a 6π-electrocyclization. A dioxenone ring-opening/acyl ketene trapping reaction with a primary amide provided the unusual unsymmetrical imide functionality. Other key steps include the use of an Evans auxiliary alkylation (d.r. ≥ 95:5) to install the (S)-2-methyl succinic acid fragment and a cross-metathesis to install the unsaturated side-chain.

Association of SIRT6 circulating levels with urinary and glycometabolic markers in pre-diabetes and diabetes.

AIM: The study is aimed to detect the expression of serum Sirtuin 6 (SIRT6) with different severities and urinary albumin creatinine ratios (UACR) in type 2 diabetes mellitus (T2DM) patients, thus exploring the association of SIRT6 together with glycolipid metabolism and urinary protein in the cross-sectional study. METHODS: T2DM patients (313 cases), pre-diabetic patients (102 cases), and healthy volunteers (100 cases) were selected. T2DM patients were divided into the normal albuminuria (103 cases, UACR < 30 mg/g), micro-albuminuria (106 cases, UACR 30-300 mg/g), and large amount of albuminuria group (104 cases, UACR > 300 mg/g) based on different UACR levels. The medical history was asked, biochemical indicators were detected, hematuria samples were taken, serum SIRT6 levels were detected, and detailed statistical analysis was conducted. RESULTS: FPG, 2 h-PG, HOMA-IR, HbA1c, and LDL-C increased, while ISI and HDL-C decreased with the aggravation of diabetic status (P < 0.05). HbA1c, UACR, TNFα, HIF1α, and SIRT6 increased with UACR in T2DM patients (P < 0.05). Correlation analysis demonstrated that SIRT6 was significantly positively correlated with glycolipid metabolism in the whole samples, and correlated with UACR, TNFα, and HIF1α in T2DM patients (P < 0.05). Ridge regression analysis showed that SIRT6 was a risk factor for both glycolipid metabolism and urinary protein (P < 0.05). CONCLUSION: SIRT6 increases with biomarkers in glycolipid metabolism and urinary protein in different severities of diabetes and UACR, which is expected to be a potential biomarker for early prediction and diagnosis related to glycolipid metabolism disorders and related nephropathy. Trial number: ChiCTR2000039808.

Pregnane X receptor (PXR) protects against cisplatin-induced acute kidney injury in mice.

Cisplatin-induced acute kidney injury (CAKI) has been recognized as one of the most serious side effects of cisplatin. Pregnane X receptor (PXR) is a ligand-dependent nuclear receptor and serves as a master regulator of xenobiotic detoxification. Increasing evidence also suggests PXR has many other functions including the regulation of cell proliferation, inflammatory response, and glucose and lipid metabolism. In this study, we aimed to investigate the role of PXR in cisplatin-induced nephrotoxicity in mice. CAKI model was performed in wild-type or PXR knockout mice. Pregnenolone 16α­carbonitrile (PCN), a mouse PXR specific agonist, was used for PXR activation. The renal function, biochemical, histopathological and molecular alterations were examined in mouse blood, urine or renal tissues. Whole transcriptome analysis was performed by RNA sequencing. We found that PXR activation significantly attenuated CAKI as reflected by improved renal function, reduced renal tubular apoptosis, ameliorated oxidative and endoplasmic reticulum stress, and suppressed inflammatory gene expression. RNA sequencing analysis revealed that the renoprotective effect of PXR was associated with multiple crucial signaling pathways, especially the PI3K/AKT pathway. In vitro study further revealed that PXR protected against cisplatin-induced apoptosis of cultured proximal tubule cells in a PI3K-dependent manner. Our results demonstrate that PXR activation can preserve renal function in cisplatin-induced AKI and suggest a possibility of PXR as a novel protective target for cisplatin-induced nephrotoxicity.

Positive association between PTN polymorphisms and schizophrenia in Northeast Chinese Han population.

As a severely and highly heritable psychotic disorder, schizophrenia has become a serious public health problem in modern society. Pleiotrophin (PTN) is a secreted cell cytokine associated with the extracellular matrix and acts as a growth factor. PTN is mainly expressed in neuroectodermal and mesodermal tissues, indicating its effect in neuron migration and epithelium-mesenchyme interactions. Whereas PTN is associated with some neurodegenerative diseases and has modulating effects on them. In this study, we aimed to investigate the association between PTN polymorphisms and schizophrenia in an independent case-control sample-set including 738 schizophrenia patients and 1085 healthy controls. Of the 13 selected single nucleotide polymorphisms (SNPs), five showed significant differences in allele or/and genotype frequencies between patients and controls: rs3959914 (genotype: χ = 11.5217, P = 0.0032); rs11765480 (genotype: χ = 10.6620, P = 0.0049); rs1473355 (genotype: χ = 8.3902, P = 0.0151); rs322246 (allele: χ = 5.5954, P = 0.0180); rs322240 (genotype: χ = 8.8429, P = 0.0121; allele: χ = 8.7802, P = 0.0031). The haplotype analysis of the selected SNPs showed different haplotype frequencies for one block (rs322240, rs322246) between cases and controls (global: χ = 9.0290, P = 0.0110; A-G: χ = 8.985, P = 0.0027; C-A: χ = 5.814, P = 0.0159). Our present results indicate PTN as a susceptibility gene for schizophrenia.

Activation of farnesoid X receptor (FXR) induces crystallin zeta expression in mouse medullary collecting duct cells.

Crystallin zeta (CRYZ) is a phylogenetically restricted water-soluble protein and provides cytoprotection against oxidative stress via multiple mechanisms. Increasing evidence suggests that CRYZ is high abundantly expressed in the kidney where it acts as a transacting factor in increasing glutaminolysis and the Na+/K+/2Cl- cotransporter (BSC1/NKCC2) expression to help maintain acid-base balance and medullary hyperosmotic gradient. However, the mechanism by which CRYZ is regulated in the kidney remains largely uncharacterized. Here, we show that CRYZ is a direct target of farnesoid X receptor (FXR), a nuclear receptor important for renal physiology. We found that CRYZ was ubiquitously expressed in mouse kidney and constitutively expressed in the cytoplasm of medullary collecting duct cells (MCDs). In primary cultured mouse MCDs, CRYZ expression was significantly upregulated by the activation and overexpression of FXR. FXR-induced CRYZ expression was almost completely abolished in the MCD cells with siRNA-mediated FXR knockdown. Consistently, treatment with FXR agonists failed to induce CRYZ expression in the MCDs isolated from mice with global and collecting duct-specific FXR deficiency. We identified a putative FXR response element (FXRE) on the CRYZ gene promoter. The luciferase reporter and ChIP assays revealed that FXR can bind directly to the FXRE site, which was further markedly enhanced by FXR activation. Furthermore, we found CRYZ overexpression in MCDs significantly attenuated hypertonicity-induced cell death possibly via increasing Bcl-2 expression. Collectively, our findings demonstrate that CRYZ is constitutively expressed in renal medullary collecting duct cells, where it is transcriptionally controlled by FXR. Given a critical role of FXR in MCDs, CRYZ may be responsible for protective effect of FXR on the survival of MCDs under hypertonic condition during dehydration.

Thioredoxin plays a key role in retinal neuropathy prior to endothelial damage in diabetic mice.

Diabetes is a chronic metabolic syndrome that results in changes in carbohydrate, lipid and protein metabolism. With diabetes for a long time, it increases the risk of diabetic retinopathy (DR) and long-term morbidity and mortality. Moreover, emerging evidence suggests that neuron damage occurs earlier than microvascular complications in DR patients, but the underlying mechanism is unclear. We investigated diabetes-induced retinal neuropathy and elucidated key molecular events to identify new therapeutic targets for the clinical treatment and prevention of DR. For in vivo studies, a high-fat diet and streptozotocin (STZ) injection were used to generate the diabetes model. Hematoxylin-eosin staining was used for morphological observations and measurements of the outer nuclear layer thickness. Electroretinography (ERG) was used to assess retinal function. For in vitro studies, Neuro2a cells were incubated in normal (5.5 mM) and high-glucose (30 mM) conditions. Flow cytometry assays were performed to analyze apoptosis. Additionally, real-time PCR and Western blotting analyses were carried out to determine gene and protein expression in vitro and in vivo. Taken together, the results indicated that retinal neuropathy occurred prior to endothelial damage induced by diabetes, and thioredoxin (Trx) plays a key role in this process. This underlying mechanism may be related to activation of the Trx/ASK1/p-p38/Trx-interacting protein pathway.

Thioredoxin is implicated in the anti­apoptotic effects of grape seed proanthocyanidin extract during hyperglycemia.

Diabetic retinopathy has long been recognized as a microvascular disease, however, recent research has indicated that diabetic retinopathy may also be considered a neurodegenerative disease. The elucidation of the molecular mechanisms underlying the development of diabetic retinopathy is imperative for the development of preventive and treatment strategies for patients with diabetes. In the present study, grape seed proanthocyanidin extract (GSPE) was used to upregulate the expression of thioredoxin (Trx), in order to evaluate its potential as a novel agent for the prevention and treatment of neurodegenerative diseases, including diabetic retinopathy. Hematoxylin and eosin staining was performed to observe the morphology of retinal neurons, whereas flow cytometry and terminal deoxynucleotidyl transferase 2'­deoxyuridine, 5'­triphosphate nick­end labeling were employed to investigate cellular apoptosis. Reverse transcription­quantitative polymerase chain reaction and western blot analysis were performed to assess the mRNA and protein expression of target proteins in order to investigate the underlying molecular mechanisms. In vivo, it was found that the photoreceptor cell was damaged in diabetic mice but following GSPE treatment, the process could be inhibited. In vitro, the results of the current study demonstrated that, under hyperglycemic culture conditions, the expression of 78 kDa glucose­regulated protein, which is an endoplasmic reticulum stress marker, was upregulated. In addition, the expression of Trx was downregulated and cell apoptosis was enhanced. Notably, treatment with GSPE was revealed to inhibit the neurodegenerative process induced by hyperglycemia. However, treatment with the Trx inhibitor PX12 in combination with GSPE was demonstrated to potentiate apoptosis compared with GSPE treatment alone under hyperglycemic conditions. Furthermore, the protein expression of apoptosis signal­regulating kinase (ASK) 1 and Trx­interacting protein (Txnip) was also upregulated by hyperglycemia, whereas GSPE was revealed to counteract this upregulation. In conclusion, the results of the present study indicate that Trx may be implicated in the mechanisms underlying the protective effects of GSPE against hyperglycemia­induced cell degeneration and apoptosis. The molecular mechanisms may also involve inhibition of the activation of the Trx/ASK1/Txnip signaling pathway.