UBIAD1 effectively alleviated myocardial ischemia reperfusion injury by activating SIRT1/PGC1α.

BACKGROUND AND OBJECTIVES: Myocardial ischemia-reperfusion injury (MIRI) threatens global health and lowers people's sense of happiness. Till now, the mechanism of MIRI has not been well-understood. Therefore, this study was designed to explore the role of UBIAD1 in MIRI as well as its detailed reaction mechanism. METHODS: The mRNA and protein expressions of UBIAD1 before or after transfection were measured using RT-qPCR and western blot. Western blot was also adopted to measure the expressions of signaling pathway-, mitochondrial damage- and apoptosis-related proteins. Moreover, mitochondrial membrane potential and ATP level were verified by JC-1 immunofluorescence and ATP kits, respectively. With the application of CCK-8, LDH and CK-MB assays, the cell viability, LDH and CK-MB levels were evaluated, respectively. In addition, the cell apoptosis was detected using TUNEL. Finally, the expressions of ROS, SOD, MDA and CAT were measured using DCFH-DA, SOD, MDA and CAT assays, respectively. RESULTS: In the present study, we found that UBIAD1 was downregulated in hypoxia-reoxygenation (H/R) -induced H9C2 cells and its upregulation could activate SIRT1/PGC1α signaling pathway. It was also found that UBIAD1 regulated mitochondrial membrane potential and ATP level via activating SIRT1/PGC1α signaling pathway. In addition, the injury of H/R-induced H9C2 cells could be relieved by UBIAD1 through the activation of SIRT1/PGC1α signaling pathway. Moreover, UBIAD1 exhibited inhibitory effects on apoptosis and oxidative stress of H/R-induced H9C2 cells through activating SIRT1/PGC1α signaling pathway. CONCLUSION: To sum up, UBIAD1 could alleviate apoptosis, oxidative stress and H9C2 cell injury by activating SIRT1/PGC1α, which laid experimental foundation for the clinical treatment of MIRI.

Pseudomonas phragmitis sp. nov., isolated from petroleum polluted river sediment.

A Gram-stain-negative, rod-shaped bacterium, motile by means of a single polar flagellum, designated S-6-2T, was isolated from petroleum polluted river sediment in Huangdao, Shandong Province, PR China. The 16S rRNA gene sequence analysis revealed that S-6-2T represented a member of the genus Pseudomonas, sharing the highest sequence similarities with Pseudomonas parafulva (97.5 %) and Pseudomonas fulva (97.5 %). Phylogenetic analysis based on 16S rRNA gene, concatenated 16S rRNA, gyrB, rpoB and rpoD genes and genome core-genes indicated that S-6-2T was affiliated with the members of the Pseudomonas pertucinogena group. The average nucleotide identity (ANI) and genome-to-genome distance between the whole genome sequences of S-6-2T and closely related species of the genus Pseudomonas within the P. pertucinogena group were less than 77.94 % and 20.5 %, respectively. Differences in phenotypic characteristics were also found between S-6-2T and the closely related species. The major cellular fatty acids (>10 %) were summed feature 8 (C18 : 1ω7c/ C18  : 1ω6c), C16 : 0, C17 : 0cyclo and C12 : 0. The predominant respiratory quinone was ubiquinone 9. The major polar lipids were diphosphatidylglycerol (DPG), phosphatidylglycerol (PG), phosphatidylethanolamine (PE), one unidentified lipid (L1), two unidentified phospholipids (PL1 and PL2) and an aminophospholipid (APL). The DNA G+C content of the genome of S-6-2T was 60.1 mol%. On the basis of the evidence from the polyphasic taxonomic study, strain S-6-2T can be classified as representative of a novel species of the genus Pseudomonas, for which the name Pseudomonas phragmitis sp. nov. is proposed. The type strain is S-6-2T (=CGMCC 1.15798T=KCTC 52539T).

Very Bright Phycoerythrobilin Chromophore for Fluorescence Biolabeling.

Biliproteins have extended the spectral range of fluorescent proteins into the far-red (FR) and near-infrared (NIR) regions. These FR and NIR fluorescent proteins are suitable for the bioimaging of mammalian tissues and are indispensable for multiplex labeling. Their application, however, presents considerable challenges in increasing their brightness, while maintaining emission in FR regions and oligomerization of monomers. Two fluorescent biliprotein triads, termed BDFP1.2/1.6:3.3:1.2/1.6, are reported. In mammalian cells, these triads not only have extremely high brightness in the FR region, but also have monomeric oligomerization. The BDFP1.2 and BDFP1.6 domains covalently bind to biliverdin, which is accessible in most cells. The BDFP3.3 domain noncovalently binds phycoerythrobilin that is added externally. A new method of replacing phycoerythrobilin with proteolytically digested BDFP3.3 facilitates this labeling. BDFP3.3 has a very high fluorescence quantum yield of 66 %, with maximal absorbance at λ=608 nm and fluorescence at λ=619 nm. In BDFP1.2/1.6:3.3:1.2/1.6, the excitation energy that is absorbed in the red region by phycoerythrobilin in the BDFP3.3 domain is transferred to biliverdin in the two BDFP1.2 or BDFP1.6 domains and fluoresces at λ≈670 nm. The combination of BDFP3.3 and BDFP1.2/1.6:3.3:1.2/1.6 can realize dual-color labeling. Labeling various proteins by fusion to these new fluorescent biliproteins is demonstrated in prokaryotic and mammalian cells.

Far-red light photoacclimation: Chromophorylation of FR induced α- and ß-subunits of allophycocyanin from Chroococcidiopsis thermalis sp. PCC7203.

Cyanobacterial light-harvesting complexes, phycobilisomes, can undergo extensive remodeling under varying light conditions. Acclimation to far-red light involves not only generation of red-shifted chlorophylls in the photosystems, but also induction of additional copies of core biliproteins that have been related to red-shifted components of the phycobilisome (Gan et al., Life 5, 4, 2015). We are studying the molecular basis for these acclimations in Chroococcidiopsis thermalis sp. PCC7203. Five far-red induced allophycocyanin subunits (ApcA2, ApcA3, ApcB2, ApcB3 and ApcF2) were expressed in Escherichia coli, together with S-type chromophore-protein lyases and in situ generated chromophore, phycocyanobilin. Only one subunit, ApcF2, shows an unusual red-shift (λAmax~675nm, λFmax~698nm): it binds the chromophore non-covalently, thereby preserving its full conjugation length. This mechanism operates also in two Cys-variants of the induced subunits of bulky APC. All other wild-type subunits bind phycocyanobilin covalently to the conventional Cys-81 under catalysis of the lyase, CpcS1. Although three of them also show binding to additional cysteines, all absorb and fluoresce similar to conventional APC subunits (λAmax~610nm, λFmax~640nm). Another origin of red-shifted complexes was identified, however, when different wild-type α- and ß-subunits of the far-red induced bulky APC were combined in a combinatorial fashion. Strongly red-shifted complexes (λFmax≤722nm) were formed when the α-subunit, PCB-ApcA2, and the ß-subunit, PCB-ApcB2, were generated together in E. coli. This extreme aggregation-induced red-shift of ~90nm of covalently bound chromophores is reminiscent, but much larger, than the ~30nm observed with conventional APC.

Structure and mechanism of the phycobiliprotein lyase CpcT.

Pigmentation of light-harvesting phycobiliproteins of cyanobacteria requires covalent attachment of open-chain tetrapyrroles, bilins, to the apoproteins. Thioether formation via addition of a cysteine residue to the 3-ethylidene substituent of bilins is mediated by lyases. T-type lyases are responsible for attachment to Cys-155 of phycobiliprotein ß-subunits. We present crystal structures of CpcT (All5339) from Nostoc (Anabaena) sp. PCC 7120 and its complex with phycocyanobilin at 1.95 and 2.50 Å resolution, respectively. CpcT forms a dimer and adopts a calyx-shaped ß-barrel fold. Although the overall structure of CpcT is largely retained upon chromophore binding, arginine residues at the opening of the binding pocket undergo major rotameric rearrangements anchoring the propionate groups of phycocyanobilin. Based on the structure and mutational analysis, a reaction mechanism is proposed that accounts for chromophore stabilization and regio- and stereospecificity of the addition reaction. At the dimer interface, a loop extending from one subunit partially shields the opening of the phycocyanobilin binding pocket in the other subunit. Deletion of the loop or disruptions of the dimer interface significantly reduce CpcT lyase activity, suggesting functional relevance of the dimer. Dimerization is further enhanced by chromophore binding. The chromophore is largely buried in the dimer, but in the monomer, the 3-ethylidene group is accessible for the apophycobiliprotein, preferentially from the chromophore α-side. Asp-163 and Tyr-65 at the ß- and α-face near the E-configured ethylidene group, respectively, support the acid-catalyzed nucleophilic Michael addition of cysteine 155 of the apoprotein to an N-acylimmonium intermediate proposed by Grubmayr and Wagner (Grubmayr, K., and Wagner, U. G. (1988) Monatsh. Chem. 119, 965-983).

The applied anatomy of anterior approach for minimally invasive hip joint surgery.

The anterior approach for minimally invasive hip joint surgery is one of the common approaches utilized in hip joint surgery. Here, we report the results of dissections in 60 sides of human adult cadavers. We observed and measured the branches of the superficial circumflex iliac artery, the lateral femoral cutaneous nerves, the lateral circumflex femoral artery, and the superior gluteal nerves in the experiment via the anterior approach for minimally invasive hip joint surgery. The relationship between these structures and the anterior approach was studied. The present study provides important data demonstrating the location, path of dominant structures that might be encountered during the surgery and their relationships with the surgical incision. These data may allow surgeons performing the anterior approach for hip joint surgery to minimize the risk of neurovascular injury.

[Understanding the planes of total mesorectal excision through surgical anatomy of pelvic fascia].

OBJECTIVE: To study the relationship of mesorectum with fasciae and nerves in the pelvic cavity and to specify the proper planes of dissection in total mesorectal excision. METHODS: Twenty-four pelvises (12 males and 12 females) harvested from cadavers were studied by dissection. RESULTS: There were three planes surrounding the rectum as the visceral fascia, vesicohypogastric fascia and parietal fascia. The pelvic plexus and its branches situated between the visceral fascia and the vesicohypogastric fascia. Pelvic splanchnic nerves and hypogastric nerves were observed between the visceral fascia and the parietal fascia. CONCLUSIONS: The posterior plane of total mesorectal excision lies between the visceral fascia and the parietal fascia. The lateral dissection should be conducted in a plane between the visceral fascia and the vesicohypogastric fascia. The proper planes for posterior and lateral resection can be identified by the hypogastric nerve and the pelvic plexus respectively.