Prolyl Hydroxylase 3 (PHD3) Modulates Catabolic Effects of Tumor Necrosis Factor-α (TNF-α) on Cells of the Nucleus Pulposus through Co-activation of Nuclear Factor κB (NF-κB)/p65 Signaling

Recent studies suggest a differential role of prolyl hydroxylase (PHD) isoforms in controlling hypoxia-inducible factor (HIF)-α degradation and activity in nucleus pulposus (NP) cells. However, the regulation and function of PHDs under inflammatory conditions that characterize disc disease are not yet known. Here, we show that in NP cells, TNF-α and IL-1β induce PHD3 expression through NF-κB. Lentiviral delivery of Sh-p65 and Sh-IKKβ confirms that cytokine-mediated PHD3 expression is NF-κB-dependent. It is noteworthy that although both cytokines induce HIF activity, mechanistic studies using Sh-HIF-1α and PHD3 promoter/enhancer constructs harboring well characterized hypoxia response element (HRE) show lack of HIF involvement in cytokine-mediated PHD3 expression. Loss-of-function studies clearly indicate that PHD3 serves as a co-activator of NF-κB signaling activity in NP cells; PHD3 interacts with, and co-localizes with, p65. We observed that when PHD3 is silenced, there is a significant decrease in TNF-α-induced expression of catabolic markers that include ADAMTS5, syndecan4, MMP13, and COX2, and at the same time, there is restoration of aggrecan and collagen type II expression. It is noteworthy that hydroxylase function of PHDs is not required for mediating cytokine-dependent gene expression. These findings show that by enhancing the activity of inflammatory cytokines, PHD3 may serve a critical role in degenerative disc disease.     

Crystal structure of the C-terminal domain of Mu phage central spike and functions of bound calcium ion

Bacteriophage Mu, which has a contractile tail, is one of the most famous genus of Myoviridae. It has a wide host range and is thought to contribute to horizontal gene transfer. The Myoviridae infection process is initiated by adhesion to the host surface. The phage then penetrates the host cell membrane using its tail to inject its genetic material into the host. In this penetration process, Myoviridae phages are proposed to puncture the membrane of the host cell using a central spike located beneath its baseplate. The central spike of the Mu phage is thought to be composed of gene 45 product (gp45), which has a significant sequence homology with the central spike of P2 phage (gpV). We determined the crystal structure of shortened Mu gp45Δ1-91 (Arg92–Gln197) at 1.5 Å resolution and showed that Mu gp45 is a needlelike structure that punctures the membrane. The apex of Mu gp45 and that of P2 gpV contained iron, chloride, and calcium ions. Although the C-terminal domain of Mu gp45 was sufficient for binding to the E. coli membrane, a mutant D188A, in which the Asp amino acid residue that coordinates the calcium ion was replaced by Ala, did not exhibit a propensity to bind to the membrane. Therefore, we concluded that calcium ion played an important role in interaction with the host cell membrane.     

Impact of long-term continuous positive airway pressure on liver fat in male obstructive sleep apnea patients with fatty liver

Sleep and Biological Rhythms - Obstructive sleep apnea (OSA) is an established factor in the pathogenesis and exacerbation of fatty liver disease. However, randomized controlled trials have failed...     

Metabolism of14C-labeled epibrassinolide in intact seedlings of cucumber and wheat

The metabolism of exogenously applied¹⁴C-24-epibrassinolide (¹⁴C-EBR) in seedlings of cucumber and wheat was examined. Total lipids were extracted with isopropanol and chloroform, and then partitioned with water. More than 80% of radioactivity was distributed to chloroform-phase. Concentrated chloroform-phase was applied to silica gel plate and was developed with chloroform-ethanol (5:1 v/v). Rf value of original¹⁴C-EBR was about 0.6. In cucumber leaves harvested after 2 day-culture, three peaks were detected at Rf 0.11, 0.47 and 0.84. In cucumber petioles of 2 day-culture, however, a major peak was detected at Rf 0.90. But¹⁴C-EBR was hardly metabolized in hypocotyls and roots after 2 days. In wheat leaves harvested just after pulse labeling, a peak was detected at Rf 0.63. By further analysis of this peak using ODS-HPLC, however, an original peak of¹⁴C-EBR and two metabolites having higher polarity were detected. In wheat leaves harvested after 2 day-culture, the profile of TLC scanning was similar to that just after pulse labeling, although, an original peak of¹⁴C-EBR was no longer detected by ODS-HPLC. In wheat roots,¹⁴C-EBR was hardly metabolized. These results indicate that¹⁴C-EBR occurring in leaves and petioles is metabolized to produce several kinds of metabolites.     

Selective Estrogen Receptor Modulators Suppress Hif1α Protein Accumulation in Mouse Osteoclasts

Anti-bone resorptive drugs such as bisphosphonates, the anti-RANKL antibody (denosumab), or selective estrogen receptor modulators (SERMs) have been developed to treat osteoporosis. Mechanisms underlying activity of bisphosphonates or denosumab in this context are understood, while it is less clear how SERMs like tamoxifen, raloxifene, or bazedoxifene inhibit bone resorption. Recently, accumulation of hypoxia inducible factor 1 alpha (Hif1α) in osteoclasts was shown to be suppressed by estrogen in normal cells. In addition, osteoclast activation and decreased bone mass seen in estrogen-deficient conditions was found to require Hif1α. Here, we used western blot analysis of cultured osteoclast precursor cells to show that tamoxifen, raloxifene, or bazedoxifene all suppress Hif1α protein accumulation. The effects of each SERM on osteoclast differentiation differed in vitro. Our results suggest that interventions such as the SERMs evaluated here could be useful to inhibit Hif1α and osteoclast activity under estrogen-deficient conditions.     

Ultrastructural study of crystalloids in Sertoli cells of the three-toed sloth (Bradypus tridactylus)

Summary Crystalloids were found in Sertoli cells of the testis of the three-toed sloth by examination at the lightand electron-microscopic levels. Needle-, or spindle-shaped crystalloids, varying in length, were located in the basal part of the Sertoli cells. They consisted of bundles of filaments each measuring ~ 11 nm in diameter. Several filaments were packed hexagonally to form a bundle. The center-to-center distance between individual filaments of a bundle was ~ 17 nm. Periodical lateral projections emanated from the filaments. Cross sections of crystalloids showed that the projections radiated from each filament in three directions, forming an equilateral triangle with a side length of ~ 15 nm. Scattered polyribosomes were found between and around the bundles.     

Effects of antibiotics on the chloroplast replication in the mossPlagiomnium trichomanes

Protoplasma - Four morphological processes were observed during the cycle of the chloroplast replication of the mossPlagiomnium trichomanes; spherical to ellipsoidal, ellipsoidal to slight...     

Biochemical polymorphism in the rat,Rattus norvegicus: Genetic study of four markers

The linkage of the hemoglobin (Hbb) and albino (c) loci has been determined from backcross progeny of the mating (WAG/Orl × Long Evans/Orl)F ₁ × WAG/Orl. The data give 9.1 ± 1.8% recombination. The backcross (August/Orl × WAG/Orl)F ₁ × August/Orl segregating for Hbb and pink-eyed yellow (p) shows 21.5±4.2% recombination. The proposed gene order on linkage group I is p-c-Hbb. Linkage of the seminal vesicle protein (Svp-1) and the nonagouti (a) loci has been determined from backcross progeny of the mating (August/Orl × Long Evans/Orl)F ₁ × Long Evans/Orl. The data show 7.1±3.4% recombination. Svp-1 thus represents an additional marker in linkage group V. Two new autosomal variants have been reported: The locus which controls a plasma protein's polymorphism has been designated Gl-1 with two codominant alleles Gl-1a and Gl-1b. The other locus, controlling a testis esterases polymorphism, has been assigned the symbol Es-3 and has two codominant alleles Es-3a and Es-3b. The absence of linkage of Gl-1 and Es-3 to each other and to five different loci has also been reported. Rat and mouse analogy with respect to these markers and established linkages is discussed.