The Angiopoietin-1 Variant COMP-Ang1 Enhances BMP2-Induced Bone Regeneration with Recruiting Pericytes in Critical Sized Calvarial Defects

Craniofacial bone defects are observed in a variety of clinical situations, and their reconstructions require coordinated coupling between angiogenesis and osteogenesis. In this study, we explored the effects of cartilage oligomeric matrix protein-angiopoietin 1 (COMP-Ang1), a synthetic and soluble variant of angiopoietin 1, on bone morphogenetic protein 2 (BMP2)-induced cranial bone regeneration, and recruitment and osteogenic differentiation of perivascular pericytes. A critical-size calvarial defect was created in the C57BL/6 mouse and COMP-Ang1 and/or BMP2 proteins were delivered into the defects with absorbable collagen sponges. After 3 weeks, bone regeneration was evaluated using micro-computed tomography and histologic examination. Pericyte recruitment into the defects was examined using immunofluorescence staining with anti-NG2 and anti-CD31 antibodies. In vitro recruitment and osteoblastic differentiation of pericyte cells were assessed with Boyden chamber assay, staining of calcified nodules, RT-PCR and Western blot analyses. Combined administration of COMP-Ang1 and BMP2 synergistically enhanced bone repair along with the increased population of CD31 (an endothelial cell marker) and NG2 (a specific marker of pericyte) positive cells. In vitro cultures of pericytes consistently showed that pericyte infiltration into the membrane pore of Boyden chamber was more enhanced by the combination treatment. In addition, the combination further increased the osteoblast-specific gene expression, including bone sialoprotein (BSP), osteocalcin (OCN) and osterix (OSX), phosphorylation of Smad/1/5/8, and mineralized nodule formation. COMP-Ang1 can enhance BMP2-induced cranial bone regeneration with increased pericyte recruitment. Combined delivery of the proteins might be a therapeutic strategy to repair cranial bone damage.     

The cochlea of the enigmatic pygmy right whale Caperea marginata informs mysticete phylogeny

The pygmy right whale, Caperea marginata , is the least understood extant baleen whale (Cetacea, Mysticeti). Knowledge on its basic anatomy, ecology, and fossil record is limited, even though its singular position outside both balaenids (right whales) and balaenopteroids (rorquals + grey whales) gives Caperea a pivotal role in mysticete evolution. Recent investigations of the cetacean cochlea have provided new insights into sensory capabilities and phylogeny. Here, we extend this advance to Caperea by describing, for the first time, the inner ear of this enigmatic species. The cochlea is large and appears to be sensitive to low‐frequency sounds, but its hearing limit is relatively high. The presence of a well‐developed tympanal recess links Caperea with cetotheriids and balaenopteroids, rather than balaenids, contrary to the traditional morphological view of a close Caperea‐balaenid relationship. Nevertheless, a broader sample of the cetotheriid Herpetocetus demonstrates that the presence of a tympanal recess can be variable at the specific and possibly even the intraspecific level.     

Control of the pericentrosomal H2O2 level by peroxiredoxin I is critical for mitotic progression

Proteins associated with the centrosome play key roles in mitotic progression in mammalian cells. The activity of Cdk1-opposing phosphatases at the centrosome must be inhibited during early mitosis to prevent premature dephosphorylation of Cdh1—an activator of the ubiquitin ligase anaphase-promoting complex/cyclosome—and the consequent premature degradation of mitotic activators. In this paper, we show that reversible oxidative inactivation of centrosome-bound protein phosphatases such as Cdc14B by H₂O₂ is likely responsible for this inhibition. The intracellular concentration of H₂O₂ increases as the cell cycle progresses. Whereas the centrosome is shielded from H₂O₂ through its association with the H₂O₂-eliminating enzyme peroxiredoxin I (PrxI) during interphase, the centrosome-associated PrxI is selectively inactivated through phosphorylation by Cdk1 during early mitosis, thereby exposing the centrosome to H₂O₂ and facilitating inactivation of centrosome-bound phosphatases. Dephosphorylation of PrxI by okadaic acid–sensitive phosphatases during late mitosis again shields the centrosome from H₂O₂ and thereby allows the reactivation of Cdk1-opposing phosphatases at the organelle.     

Relationship between Initial Telomere Length,Initial Telomerase Activity,Age, and Replicative Capacity of Nucleus Pulposus Chondrocytes in Human Intervertebral Discs: What Is a Predictor of Replicative Potential?

There is evidence that telomere length (TL), telomerase activity (TA), and age are related to the replicative potential of human nucleus pulposus chondrocytes (NPCs). However, it has not yet been established if any of these factors can serve as predictors of the replicative potential of NPCs. To establish predictors of the replicative potential of NPCs, we evaluated potential relationships between replicative capacity of NPCs, initial TL (telomere length at the first passage), initial TA (telomerase activity at the first passage), and age. Nucleus pulposus specimens were obtained from 14 patients of various ages undergoing discectomy. NPCs were serially cultivated until the end of their replicative lifespans. Relationships among cumulative population doubling level (PDL), initial TL, initial TA, and age were analyzed. Initial TA was negatively correlated with age (r = -0.674, P = 0.008). However, no correlation between initial TL and age was observed. Cumulative PDL was also negatively correlated with age (r = -0.585, P = 0.028). Although the cumulative PDL appeared to increase with initial TL or initial TA, this trend was not statistically significant. In conclusion, age is the sole predictor of the replicative potential of human NPCs, and replicative potential decreases with age. Initial TL and initial TA are not predictors of replicative potential, and can serve only as reference values.     

Expression of mosquitocidal crystal protein genes in non-insecticidal Bacillus thuringiensis subsp. israelensis

Bacillus thuringiensis NTB-1 isolated from soil samples in Korea produces ovoidal parasporal inclusions with proteins of approximately 24–40 kDa in size. Although serological study indicated that the isolate has a flagella (H) antigen identical with subsp. israelensis , it seemed to be non-insecticidal against Lepidoptera and Coleoptera as well as Diptera. To investigate the activity of non-insecticidal B. thuringiensis transformed with insecticidal crystal protein genes, cryIVD and cytA genes of B. thuringiensis subsp. morrisoni PG-14, highly toxic to mosquito larvae, were introduced into the isolate NTB-1. The expression of mosquitocidal crystal protein genes in NTB-1 was characterized by SDS–PAGE analysis and electron microscopy. The results showed that crystalline inclusions of host, CryIVD and CytA were stably expressed in the transformant. However, the mosquitocidal activity of transformant was similar to that of B. thuringiensis subsp. kurstaki Cry⁻ B harbouring cryIVD and cytA genes, demonstrating that a synergistic effect by an interaction of both introduced insecticidal and resident non-insecticidal crystal proteins was not observed.     

Modulation of Intracellular Calcium Levels by Calcium Lactate Affects Colon Cancer Cell Motility through Calcium-Dependent Calpain

Cancer cell motility is a key phenomenon regulating invasion and metastasis. Focal adhesion kinase (FAK) plays a major role in cellular adhesion and metastasis of various cancers. The relationship between dietary supplementation of calcium and colon cancer has been extensively investigated. However, the effect of calcium (Ca²⁺) supplementation on calpain-FAK-motility is not clearly understood. We sought to identify the mechanism of FAK cleavage through Ca²⁺ bound lactate (CaLa), its downstream signaling and role in the motility of human colon cancer cells. We found that treating HCT116 and HT-29 cells with CaLa immediately increased the intracellular Ca²⁺ (iCa²⁺) levels for a prolonged period of time. Ca²⁺ influx induced cleavage of FAK into an N-terminal FAK (FERM domain) in a dose-dependent manner. Phosphorylated FAK (p-FAK) was also cleaved in to its p-N-terminal FAK. CaLa increased colon cancer cells motility. Calpeptin, a calpain inhibitor, reversed the effects of CaLa on FAK and pFAK cleavage in both cancer cell lines. The cleaved FAK translocates into the nucleus and modulates p53 stability through MDM2-associated ubiquitination. CaLa-induced Ca²⁺ influx increased the motility of colon cancer cells was mediated by calpain activity through FAK and pFAK protein destabilization. In conclusion, these results suggest that careful consideration may be given in deciding dietary Ca²⁺ supplementation to patient undergoing treatment for metastatic cancer.     

Physiological functions of the TRPM4 channels via protein interactions

Transient Receptor Potential, Melastatin-related, member 4 (TRPM4) channels are Ca²⁺-activated Ca²⁺-impermeable cation channels. These channels are expressed in various types of mammalian tissues including the brain and are implicated in many diverse physiological and pathophysiological conditions. In the past several years, the trafficking processes and regulatory mechanism of these channels and their interacting proteins have been uncovered. Here in this minireview, we summarize the current understanding of the trafficking mechanism of TRPM4 channels on the plasma membrane as well as heteromeric complex formation via protein interactions. We also describe physiological implications of protein-TRPM4 interactions and suggest TRPM4 channels as therapeutic targets in many related diseases. [BMB Reports 2015; 48(1): 1-5]     

A comparison between biochemical tests and the 'CAMP' reaction for identification of Aeromonas spp.

Aeromonas bacteria (110 strains) from a variety of clinical, food and environmental sources, were identified using routine biochemical tests. Concurrently they were tested aerobically and anaerobically for their ability to perform synergistic haemolysis with Staphylococcus aureus (the 'CAMP' reaction). Results did not support a reported observation that the 'CAMP' reaction can he used to facilitate speciation of Aeromonas bacteria.