Sialyl alpha(2-->3) lactose clusters using carbosilane dendrimer core scaffolds as influenza hemagglutinin blockers

An efficient synthesis of a series of carbosilane dendrimers uniformly functionalized with sialyl α(2 → 3) lactose (Neu5Acα(2 → 3)Galβ(1 → 4)Glcβ1→) moieties was accomplished. The results of a preliminary study on biological responses against influenza virus hemagglutinin, using the sialyl lactose clusters showed unique biological activities on the basis of the structure–activity relationship according to the carbosilane scaffolds.     

RB1CC1 together with RB1 and p53 predicts long-term survival in Japanese breast cancer patients

RB1-inducible coiled-coil 1 (RB1CC1) plays a significant role in the enhancement of the retinoblastoma tumor suppressor (RB1) pathway and is involved in breast cancer development. However, RB1CC1's role in clinical progression of breast cancer has not yet been evaluated, so, as a first step, it is necessary to establish its usefulness as a tool to evaluate breast cancer patients. In this report, we have analyzed the correlation between abnormalities in the RB1CC1 pathway and long-term prognosis, because disease-specific death in later periods (>5 years) of the disease is a serious problem in breast cancer. Breast cancer tissues from a large cohort in Japan were evaluated by conventional immunohistochemical methods for the presence of the molecules involved in the RB1CC1 pathway, including RB1CC1, RB1, p53, and other well-known prognostic markers for breast cancer, such as estrogen receptor, progesterone receptor, and human epidermal growth factor receptor 2. The correlation between the immunohistochemical results and clinical outcomes of 323 breast cancer patients was analyzed using a Kaplan-Meier log-rank test and a multivariate Cox proportional hazards regression analysis. Absence of nuclear RB1CC1 expression was associated with the worst prognosis (Log-rank test, Chi-Square value = 17.462, p<0.0001). Dysfunction of either one of RB1CC1, RB1, or p53 was associated with the highest risk for cancer-specific death, especially related to survival lasting more than 5 years (multivariate Cox proportional hazard ratio = 3.951, 95% Confidence Interval =1.566-9.967, p = 0.0036). Our present data demonstrate that the combined evaluation of RB1CC1, RB1 and p53 by conventional immunohistochemical analysis provides an accurate prediction of the long-term prognoses of breast cancer patients, which can be carried out as a routine clinical examination.     

PINK1 is degraded through the N-end rule pathway

PINK1, a mitochondrial serine/threonine kinase, is the product of a gene mutated in an autosomal recessive form of Parkinson disease. PINK1 is constitutively degraded by an unknown mechanism and stabilized selectively on damaged mitochondria where it can recruit the E3 ligase PARK2/PARKIN to induce mitophagy. Here, we show that, under steady-state conditions, endogenous PINK1 is constitutively and rapidly degraded by E3 ubiquitin ligases UBR1, UBR2 and UBR4 through the N-end rule pathway. Following precursor import into mitochondria, PINK1 is cleaved in the transmembrane segment by a mitochondrial intramembrane protease PARL generating an N-terminal destabilizing amino acid and then retrotranslocates from mitochondria to the cytosol for N-end recognition and proteasomal degradation. Thus, sequential actions of mitochondrial import, PARL-processing, retrotranslocation and recognition by N-end rule E3 enzymes for the ubiquitin proteosomal degradation defines the rapid PINK1 turnover. PINK1 steady-state elimination by the N-end rule identifies a novel organelle to cytoplasm turnover pathway that yields a mechanism to flag damaged mitochondria for autophagic elimination.     

The Src tyrosine kinase pathway regulates thecal CYP17 expression and androstenedione secretion

In order to evaluate the role of Src tyrosine kinase in thecal cell steroidogenesis, a pharmacological approach was utilized by treating enriched populations of mouse ovarian theca-interstitial cells in vitro with a direct Src kinase inhibitor, PP2. Inhibition of Src with PP2 increased both basal and forskolin-stimulated androstenedione secretion, and increased cytochrome P450 17-alpha hydroxylase-lyase (CYP17) promoter activity and steady state mRNA. PP2 did not change thecal levels of StAR mRNA. Inhibition of mitogen-activated protein kinase kinase, a downstream regulator of Src activity, using PD98059 also increased forskolin-stimulated secretion of androstenedione above forskolin alone, but had no effect on basal secretion of androstenedione. Src inhibition increased mitogen-activated protein kinase phosphatase-1 protein and decreased phosphorylation of SF-1, which correlated with increased CYP17 promoter activity and mRNA levels. These results implicate Src tyrosine kinase in the regulation of CYP17 and thecal androgen secretion.     

Early activation does not translate into effector differentiation of peripheral CD8T cells during the acute phase of Kawasaki disease

Early diagnosis of acute Kawasaki disease (KD), lying in the spectrum between infectious and autoimmune diseases, can be difficult. To clarify the role of peripheral CD8T cells in KD, we examined their activation, proliferation, maturation, and effector function by four-color flow cytometry. Compared to healthy/febrile controls, acute KD patients showed striking increase in early activation marker CD69⁺CD8T cells and maturation subsets, but HLA-DR⁺CD8T cells representing late activation did not increase. Although Ki67⁺CD8T cells reflecting ongoing cell division increased in acute KD and febrile controls, absolute numbers of CD8T cells and maturation subsets decreased in acute KD versus healthy controls. Effector cells were lower in acute than in convalescent KD. Perforin⁺CD8T cells, denoting cytolytic activity, were lower in KD patients versus febrile controls. CD69⁺CD8T cells increase in acute KD but effector differentiation is absent. CD69⁺CD8T cells could be a marker to determine disease progression, treatment response, and convalescence in acute KD.     

Enzymatic synthesis of unique sialyloligosaccharides using marine bacterial α-(2→3)- and α-(2→6)-sialyltransferases

We investigated the acceptor substrate specificities of marine bacterial α-(2→3)-sialyltransferase cloned from Photobacterium sp. JT-ISH-224 and α-(2→6)-sialyltransferase cloned from Photobacterium damselae JT0160 using several saccharides as acceptor substrates. After purifying the enzymatic reaction products, we confirmed their structure by NMR spectroscopy. The α-(2→3)-sialyltransferase transferred N-acetylneuraminic acid (Neu5Ac) from cytidine 5′-monophospho-N-acetylneuraminic acid (CMP-Neu5Ac) to the β-anomeric hydroxyl groups of mannose (Man) and α-Manp-(1→6)-Manp, and α-(2→6)-sialyltransferase transferred N-acetylneuraminic acid to the 6-OH groups of the non-reducing end galactose residues in β-Galp-(1→3)-GlcpNAc and β-Galp-(1→6)-GlcpNAc.     

Development of Implant/Interconnected Porous Hydroxyapatite Complex as New Concept Graft Material

Background

Dental implant has been successfully used to replace missing teeth. However, in some clinical situations, implant placement may be difficult because of a large bone defect. We designed novel complex biomaterial to simultaneously restore bone and place implant. This complex was incorporated implant into interconnected porous calcium hydroxyapatite (IP-CHA). We then tested this Implant/IP-CHA complex and evaluated its effect on subsequent bone regeneration and implant stability in vivo.

Methodology/Principal Findings

A cylinder-type IP-CHA was used in this study. After forming inside of the cylinder, an implant was placed inside to fabricate the Implant/IP-CHA complex. This complex was then placed into the prepared bone socket in the femur of four beagle-Labrador hybrid dogs. As a control, implants were placed directly into the femur without any bone substrate. Bone sockets were allowed to heal for 2, 3 and 6 months and implant stability quotients (ISQ) were measured. Finally, tissue blocks containing the Implant/IP-CHA complexes were harvested. Specimens were processed for histology and stained with toluidine blue and bone implant contact (BIC) was measured. The ISQs of complex groups was 77.8±2.9 in the 6-month, 72.0±5.7 in the 3-month and 47.4±11.0 in the 2-month. There was no significant difference between the 3- or 6-month complex groups and implant control groups. In the 2-month group, connective tissue, including capillary angiogenesis, was predominant around the implants, although newly formed bone could also be observed. While, in the 3 and 6-month groups, newly formed bone could be seen in contact to most of the implant surface. The BICs of complex groups was 2.18±3.77 in the 2-month, 44.03±29.58 in the 3-month, and 51.23±8.25 in the 6-month. Significant difference was detected between the 2 and 6-month.

Conclusions/Significance

Within the results of this study, the IP-CHA/implant complex might be able to achieve both bone reconstruction and implant stability.     

Structural characteristics of green tea catechins for formation of protein carbonyl in human serum albumin

Catechins are polyphenolic antioxidants found in green tea leaves. Recent studies have reported that various polyphenolic compounds, including catechins, cause protein carbonyl formation in proteins via their pro-oxidant actions. In this study, we evaluate the formation of protein carbonyl in human serum albumin (HSA) by tea catechins and investigate the relationship between catechin chemical structure and its pro-oxidant property. To assess the formation of protein carbonyl in HSA, HSA was incubated with four individual catechins under physiological conditions to generate biotin-LC-hydrazide labeled protein carbonyls. Comparison of catechins using Western blotting revealed that the formation of protein carbonyl in HSA was higher for pyrogallol-type catechins than the corresponding catechol-type catechins. In addition, the formation of protein carbonyl was also found to be higher for the catechins having a galloyl group than the corresponding catechins lacking a galloyl group. The importance of the pyrogallol structural motif in the B-ring and the galloyl group was confirmed using methylated catechins and phenolic acids. These results indicate that the most important structural element contributing to the formation of protein carbonyl in HSA by tea catechins is the pyrogallol structural motif in the B-ring, followed by the galloyl group. The oxidation stability and binding affinity of tea catechins with proteins are responsible for the formation of protein carbonyl, and consequently the difference in these properties of each catechin may contribute to the magnitude of their biological activities.