Palladium-Catalyzed C(sp3)-H Nitrooxylation of Aliphatic Carboxamides with Practical Oxidants.

Here we report the palladium-catalyzed ß-C(sp3)-H nitrooxylation of aliphatic carboxamides using a modified quinoline auxiliary. Notably, Al(NO3)3·9H2O was used as a nitrate source as well as a practical oxidant. The 5-chloro-8-aminoquinoline auxiliary was nitrated in situ during the reaction, which may enhance its directing ability and help its removal. The reaction has a broad substrate scope with a variety of aliphatic carboxamides. The multiple substituted auxiliary can be easily removed and recovered. Two C-H-insertion palladacycle intermediates were isolated and characterized to elucidate the mechanism.

ChromInst: A single cell sequencing technique to accomplish pre-implantation comprehensive chromosomal screening overnight.

Next Generation Sequencing (NGS) is a powerful tool getting into the field of clinical examination. Its preliminary application in pre-implantation comprehensive chromosomal screening (PCCS) of assisted reproduction (test-tube baby) has shown encouraging outcomes that improves the success rate of in vitro fertilization. However, the conventional NGS library construction is time consuming. In addition with the whole genome amplification (WGA) procedure in prior, makes the single cell NGS assay hardly be accomplished within an adequately short turnover time in supporting fresh embryo implantation. In this work, we established a concise single cell sequencing protocol, ChromInst, in which the single cell WGA and NGS library construction were integrated into a two-step PCR procedure of ~ 2.5hours reaction time. We then validated the feasibility of ChromInst for overnight PCCS assay by examining 14 voluntary donated embryo biopsy samples in a single sequencing run of Miseq with merely 13M reads production. The good compatibility of ChromInst with the restriction of Illumina sequencing technique along with the good library yield uniformity resulted superior data usage efficiency and reads distribution evenness that ensures precisely distinguish of 6 normal embryos from 8 abnormal one with variable chromosomal aneuploidy. The superior succinctness and effectiveness of this protocol permits its utilization in other time limited single cell NGS applications.

Ligand symmetry significantly affects spin crossover behaviour in isomeric [Fe(pybox)2]2+ complexes.

The understanding of the correlation between the spin-state behaviour and the structural features in transition-metal complexes is of pronounced importance to the design of spin crossover compounds with high performance. However, the study of the influence of ligand symmetry on the spin crossover properties is still limited due to the shortage of suitable structural systems. Herein we report the magneto-structural correlations of three mononuclear Fe(ii) isomers with respect to their ligand symmetry. In this work, two phenyl-substituted meso and optically pure pybox ligands were employed to construct meso (1), optically pure (2), and racemic (3) ligand types of [Fe(pybox)2]2+ complexes. Their magnetic susceptibilities were measured via temperature-dependent paramagnetic 1H NMR spectroscopy. We fitted the midpoint temperatures of the transition (T1/2) of 260 K for 1(ClO4), 247 K for 2(ClO4), and 281 K for 3(ClO4). The influence of structural symmetry on spin crossover was rationalized through density functional theory calculations. The optimized structures of [Fe(pybox)2]2+ complex cations show that the geometric distortion of the central FeN6 coordination sphere is mainly caused by the steric congestions between adjacent phenyl substituents. In these compounds, there is a distinct correlation that more steric congestions produce larger coordination distortion and favor the electron configuration in the high-spin state, which reflects in the increase of T1/2. Additionally, the influence of the counter anion and lattice solvent on the meso series compounds was inspected. It is revealed that multiple factors dominate the spin-state behaviour in the solid state. This work provides deep insight into the effect of ligand symmetry on the spin transition behaviour in spin crossover compounds. It demonstrates that molecular symmetry should be considered in the design of spin crossover compounds.

Antitumor effects of saikosaponin b2 on breast cancer cell proliferation and migration.

Saikosaponin b2 (SSb2) can be extracted from Bupleurum spp. roots (Radix Bupleuri), which belongs to the Umbelliferae family. The current study aimed to explore the effects of SSb2 on proliferation of breast cancer cells and to identify the mechanism by which SSb2 affects breast cancer cell migration. mRNA expression levels of STAT3 and vasodilator­stimulated phosphoprotein (VASP) were determined and increased expression was observed in 16 breast cancer tissues compared with the paracancerous tissues. MTT, wound healing, colony formation assays and western blot suggested that SSb2 inhibited MCF­7 proliferation and migration. It was further identified by western blot analysis that SSb2 treatment reduced levels of phosphorylated STAT3, VASP, matrix metallopeptidase (MMP) 2 and MMP9 in MCF­7 compared with the untreated cells. In addition, it was demonstrated that inhibition of STAT3 phosphorylation decreased VASP expression levels and induction of STAT3 phosphorylation increased VASP levels. Furthermore, it was observed that the treatment of Kunming mice with SSb2 at 30 mg/kg/day for 30 days induced no obvious changes in the liver or kidney tissues, as determined by haematoxylin and eosin staining. In conclusion, these results indicated that SSb2 may be a potential antitumor drug for the treatment of breast cancer, which acts by suppressing proliferation and migration by downregulating the STAT3 signalling pathway and inhibiting the expression of VASP, MMP2 and MMP9 expression.

Sodium butyrate-induced apoptosis and ultrastructural changes in MCF-7 breast cancer cells.

This study investigated the effects of sodium butyrate (NaB) on Michigan Cancer Foundation-7 (MCF-7) breast cancer cells and analyzed the relevant mechanism. Here, we demonstrated that a certain concentration of NaB effectively induced MCF-7 cell apoptosis. Cell counting kit-8 (CCK-8) assay was used to detect cell viability and the apoptosis rate. Western blotting was used to detect changes in the Bcl-2 expression level. We observed cell shape changes with microscopy. Immunofluorescence revealed some apoptotic nuclei. Electron microscopy revealed thick nucleoli, chromatin margination, reduced mitochondria, and dramatic vacuoles. Collectively, our findings elucidated the morphological mechanism by which NaB changed the ultrastructure of MCF-7 cells.

Tamoxifen induces hepatotoxicity and changes to hepatocyte morphology at the early stage of endocrinotherapy in mice.

Clinically, hepatotoxicity is an inevitable side effect during long-term endocrinotherapy in breast cancer patients. Various studies have reported the specific mechanism and protective methods for this long-term hepatotoxicity, however, the short-term influences of tamoxifen (TAM) on hepatocytes remain to be elucidated. The previous study investigated TAM-induced liver injury at the early stage of endocrine treatment. Mice were assigned into 2 groups: The experiment group was administrated with intraperitoneal (i.p.) injection of 6 mg/kg/day TAM for 2 weeks, and the control group was administrated with i.p. injection of physiological saline of the same dose. Body weights in each group were detected every day, and alanine aminotransferase and aspartate aminotransferase levels were measured every 3 days. Small pieces of the liver tissues were obtained and processed for protein extraction, biochemical detection and histopathological analysis 2 weeks later. The results indicated that TAM decreased the mice body weights. Morphologically, with the treatment of TAM for only 2 weeks, at the microscopic and ultrastructural levels the structure of hepatic cords became blurred in sections of the regions, although the lobules of the liver remained visible. Partially, hepatic cells were swelled in spherical shapes. Nuclei appeared to be pyknotic and exhibited uneven chromatin distribution. In addition, it was observed in the transmission electron microscopy analysis that nuclei became pyknotic and unevenly distributed. The majority of the nuclei were endowed with distinct heterochromatin and thick nucleoli. The mitochondrial cristae became vague and disorganized. Finally, western blotting was used and detected a significant increase of the caspase-3 level in the liver tissues. In conclusion, the experiments elucidated that TAM (6 mg/kg/day) would cause hepatotoxicity at the early stage of endocrine treatment in mice, and the underlying mechanism was involved with hepatocyte apoptosis.

The Ultrastructure of MCF-7 Breast Cancer Cells after Vasodilator-Stimulated Phosphoprotein Knockdown.

Inhibition of vasodilator-stimulated phosphoprotein (VASP) expression could modulate the adhesion and proliferation of breast cancer cells. However, the underlying mechanisms are not well defined. Here, we show that knockdown of the VASP changes the ultrastructure of human MCF-7 breast cancer cells. Transfection of VASP shRNA significantly lowered the expression of VASP protein in MCF-7 cells. In the shRNA-VASP group, immunofluorescence showed diminished presence of F-actin, and it was lower in the nucleus than in the cytoplasm. After VASP was inhibited, the MCF-7 cells were oval in shape with blunt lamellipodium, disappearance of the cristae of mitochondria, decreased microvilli and more vacuoles. Collectively, our findings elucidated the morphological mechanism that knockdown of the VASP changed the ultrastructure of MCF-7 cells.

Ritonavir-induced hepatotoxicity and ultrastructural changes of hepatocytes.

To investigate the effect of ritonavir on hepatocyte proliferation, we detected the change of cleaved caspase-3 expression level in the hepatocytes. Furthermore, the morphological and ultrastructural changes of hepatocytes derived from RTV-treated mice have been observed. The results showed that ritonavir can evidently inhibit hepatocyte proliferation and increase cleaved caspase-3 expression level. Under the electron microscope, chromatin margination, mitochondrial cristae disappearance, karyopyknosis and cytoplasmic vacuolization can be observed in the hepatocytes of mice treated with ritonavir. In conclusion, the mechanism of ritonavir's hepatotoxicity is that it induces apoptosis of hepatocytes via the caspase-cascade system.

The protective effect of curcumin on hepatotoxicity and ultrastructural damage induced by cisplatin.

We investigate the protective effect of curcumin (CU) on the hepatic ultrastructural damage induced by cisplatin in mice. 18 adult Kunming mice were randomly divided into normal saline (NS) group, cisplatin treatment group (CP) and CU + CP group (n = 6 for each group). Mice in control group and CP group were administered with NS (20 mL/kg/day) and CU + CP group were i.p injected with CU (200 mg/kg/day) for 10 days. Then cisplatin (50 mg/kg/day) was injected in mice of CP group and CU + CP group, while those in control group were given the same volume of NS. Five days after injection all mice were killed and liver dissected. The hepatic morphological structures were observed under light microscope and transmission electron microscope. The results indicated that CU alleviated the hepatic histopathological damages induced by cisplatin, which included declined body weight, vacuolated cytoplasm and blurred liver trabecular structure. Moreover, no hepatic ultrastructural damages were observed in the CU protective group with condensed and marginated nuclear chromatin, bile canaliculi outstreched and bile deposited.

Protective effect of astragalus polysaccharides on liver injury induced by several different chemotherapeutics in mice.

Side effects are an unavoidable consequence of chemotherapy drugs, during which liver injury often takes place. The current study was designed to investigate the protective effect of Astragalus polysaccharides (APS) against the hepatotoxicity induced by frequently-used chemical therapy agents, cyclophosphamide (CTX), docetaxel (DTX) and epirubicin (EPI)) in mice. Mice were divided into five groups, controls, low or high dose groups (DTXL, CTXL, EPIL or DTXH, CTXH, EPIH), and low or high dose chemotherapeutics+APS groups (DTXL+APS, CTXL+APS, EPIL+APS or DTXH+APS, CTXH+APS, EPIH+APS). Controls were treated with equivalent normal saline for 28 days every other day; low or high dose group were intraperitoneal (i.p) injected with low or high doses of CTX, DTX and EPI for 28 days every other day; low or high dose chemotherapeutics+APS group were separately intraperitoneal (i.p) injected with chemotherapeutics for 28 days every other day and i.p with APS (100 mg/kg) for 7 days continually from the 22th to the 28th days. The body weight, serum levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST), histopathological features, and ultrastructure morphological change of liver tissues, protein expression level of caspase-3 were estimated at different time points. With high dose treatment of DTX, CTX and EPI, weight gain was inhibited and serum levels of ALT and AST were significantly increased. Sections of liver tissue showed massive hepatotoxicity in CTXH group compared to the control group, including hepatic lobule disorder, granular and vacuolar degeneration and necrosis in hepatic cells. These changes were confirmed at ultrastructural level, including obvious pyknosis, heterochromatin aggregation, nuclear membrane resolution, and chondrosome crystal decrease. Western blotting revealed that the protein levels of caspase-3 increased in CTXH group. The low dose groups exhibited trivial hepatotoxicity. More interestingly, after 100 mg/kg APS, liver injury was redecued not only regarding serum transaminase activities (low or high dose chemotherapeutics+APS group), but also from pathological and ultrastructural changes and the protein levels of caspase-3 (CTXH+APS group). In conclusion, DTX, CTX and EPI induce liver damage in a dose dependent manner, whereas APS exerted protective effects.

A novel self-sequence reaction network involving a set of six reactions in one pot: the synthesis of substituted benzothiazoles from aromatic ketones and anilines.

Employing simple and readily available aromatic ketones and anilines as starting materials resulted in the construction of 2-acylbenzothiazoles via a novel self-sequence reaction network, which assembles six reactions in one pot. The reaction network not only supplied a novel method for constructing complex molecules but also provided a typical example for logical self-organization synthesis.

[Correlation between vaspin concentration and insulin sensitivity in the visceral adipose tissue of young obese rats].

OBJECTIVE: To investigate the correlation between visceral adipose tissue-derived serine protease inhibitor (vaspin) concentration and insulin sensitivity in the visceral adipose tissue of young obese Sprague-Dawley (SD) rats. METHODS: Twenty-four SD rats which had been weaned 3 weeks before were randomly divided into two groups (n=12 each) to receive a high-fat and normal diet. The weight and abdominal circumference (AC) of each rat were measured, the fasting plasma glucose (FPG) and fasting insulin (FINS) in blood from the angular vein were measured after 12 hours of fasting and blood glucose (BG) and insulin (INS) levels in blood from the angular vein were measured at 60 and 120 minutes after intraperitoneal injection of 50% glucose (2 g/kg). The rats were sacrificed, and their liver and visceral adipose tissue were weighed. The vaspin concentration of the visceral adipose tissue in each rat was measured using ELISA. Correlation analysis was performed on the vaspin concentration and other indices. RESULTS: Compared with the normal diet group, the high-fat diet group showed significantly higher weight, AC, weight of visceral adipose tissue, FPG, FINS, 120 minute INS level, vaspin concentration, homeostasis model assessment for insulin resistance (HOMA-IR) and homeostasis model assessment of ß cell function (HOMA-ß) (P<0.05) Insulin sensitivity index (ISI) was significantly lower (P<0.01). Vaspin concentration was positively correlated with visceral adipose tissue and liver weight, AC, 120 minute INS level, FPG, FINS, HOMA-IR and HOMA-ß (P<0.05), and negatively correlated with ISI (P<0.05). CONCLUSIONS: High expression of vaspin is associated with insulin resistance in young obese SD rats. Vaspin is presumably an adipocytokine that can increase insulin sensitivity, promote insulin secretion by islet ß-cells and improve glucose tolerance, and it may be involved in insulin resistance and the disturbance of carbohydrate metabolism.

Diversity-oriented synthesis of chromenes via metal-free domino reactions from ketones and phenols.

Functionalized chromenes have been synthesized via highly selective metal-free domino reactions from ketones and phenols. 2H-Chromenes, 4H-chromenes, spiran and benzocyclopentane can be respectively prepared starting from the corresponding cyclic ketones, aryl methyl ketones, acetone, and 3-pentanone.

[Expression of MICA/B protein in esophageal cancer and its clinical significance].

OBJECTIVE: To explore the expression of MICA/B in human esophageal cancer, and to analyze its correlation with clinicopathological features. METHODS: The expression of MICA/B in 40 cases of esophagus carcinoma and corresponding normal esophageal mucosa tissues were examined by immunohistochemistry. RESULTS: The positive rate of expression of MICA/B protein in the esophageal carcinoma was 75.0% (30/40), and that in the corresponding normal esophageal mucosa was 0 (0/40). Up-regulation of MICA/B expression was found in the esophageal carcinomas. The expression of MICA/B was related with histological grade of the esophageal carcinoma (P = 0.012). CONCLUSION: MICA/B protein plays an important role in the esophageal carcinogenesis, and my become a useful molecular marker for the diagnosis of esophageal carcinoma.