To assess the histological response and the reinforcing effects of bone ingrowth within porous hydroxyapatite (HA) implants depending on pore geometry, four kinds of cylindrical-type with parallel ...linear pores (
φ50, 100, 300, 500
μm), one kind of sponge-type with irregular interconnecting pores (
φ250
μm) and one cross-type with crossing linear pores (
φ100×120
μm) of porous HA were prepared. Eighty-four rabbits were divided into six groups, and a 5×5×7
mm sized porous HA block was inserted through the medial cortical window of the proximal tibia. Histomorphological changes were examined using light and scanning electron microscopy. A biomechanical compression test was performed using material test machines. After implantation, the implants showed different histological changes depending on pore geometry. Active osteoconduction was also found in the
φ50
μm sized cylindrical-type porous HA. Evidence of remodeling of new bone and bone marrow formation within porous HA was found in the larger cylindrical-types (
φ300, 500
μm), and the sponge- and cross-types. The biomechanical test showed that the ultimate compressive strength increased significantly in the
φ300
μm sized cylindrical-type, and in the sponge- and cross-types eight weeks after implantation. Porous HA with cylindrical pores could be a useful graft material due to its strength, osteoconductivity and the ease with which its pore geometry can be controlled.
Pulmonary lymphangioleiomyomatosis is a progressive cystic lung disease that is associated with infiltration of atypical smooth muscle-like cells. Previous descriptions of clinical characteristics of ...subjects with lymphangioleiomyomatosis have been based on a limited number of patients.
To describe the clinical characteristics of subjects with pulmonary lymphangioleiomyomatosis, both sporadic and tuberous sclerosis-related forms.
Over a 3-yr period, from 1998 to 2001, 243 subjects with pulmonary lymphangioleiomyomatosis were enrolled into a national registry; 13 subjects who had already undergone lung transplantation were excluded for the purposes of this report.
All 230 subjects were women, aged 18 to 76 yr (mean +/- SE, 44.5 +/- 0.65 yr). The average age at onset of symptoms was 38.9 +/- 0.73 yr and at diagnosis was 41.0 +/- 0.65 yr. Tuberous sclerosis complex was present in 14.8% of subjects. Pulmonary manifestations, most commonly spontaneous pneumothorax, were the primary events leading to the diagnosis in 86.5% of cases. Nearly 55% of the subjects were being treated with a progesterone derivative. An obstructive pattern on pulmonary function testing was observed in 57.3% of the subjects, whereas 33.9% had normal spirometric results. Women with tuberous sclerosis-related lymphangioleiomyomatosis were younger and had less impaired lung function compared with those with the sporadic form.
The age range of women afflicted with pulmonary lymphangioleiomyomatosis is broader than previously appreciated and the degree of pulmonary function can be quite variable, with one-third of subjects having normal spirometry at enrollment into this registry.
Mesenchymal stem cell (MSC)-based gene therapy is a promising tool for the treatment of various neurological diseases, including brain tumors. However, the tracking of in vivo stem cell migration, ...distribution, and survival need to be defined for their clinical application. The systemic routes of stem cell delivery must be determined because direct intracerebral injection as a cure for brain tumors is an invasive method. In this study, we show for the first time that near-infrared (NIR) imaging can reveal the distribution and tumor tropism of intravenously injected MSCs in an intracranial xenograft glioma model. MSCs were labeled with NIR fluorescent nanoparticles, and the effects of the NIR dye on cell proliferation and migratory capacity were evaluated in vitro. We investigated the tumor-targeting properties and tissue distribution of labeled MSCs introduced by intravenous injection and followed by in vivo imaging analysis, histological analysis, and real-time quantitative polymerase chain reaction. We observed no cytotoxicity or change in the overall growth rate and characteristics of labeled MSCs compared with control MSCs. NIR fluorescent imaging showed the organ distribution and targeted tumor tropism of systemically injected human MSCs. A significant number of MSCs accumulated specifically at the tumor site in the mouse brain. These results suggest that NIR-based cell tracking is a potentially useful imaging technique to visualize cell survival, migration, and distribution for the application of MSC-mediated therapies in the treatment of malignant gliomas.
Among other emerging amyloid-targeting blood-based biomarkers, Multimer Detection System-Oligomeric Amyloid-β (MDS-OAβ) measures dynamic changes in concentration of oligomeric amyloid-β (OAβ), which ...is considered the main pathogenic culprit of Alzheimer's disease (AD), in plasma after spiking with synthetic amyloid-β (Aβ). We aimed to investigate the predictability of MDS-OAβ on amyloid positron emission tomography (PET) positivity.
A total of 96 subjects who visited Seoul National University Bundang Hospital for medical check-up complaining of cognitive decline and had undergone extensive medical assessment were recruited. Amyloid statuses were dichotomized into positive or negative based on visual assessment of amyloid PET. Plasma OAβ concentration was measured by MDS-OAβ. In the previous validation study, 0.78ng/mL was established as the cut-off value and the plasma OAβ concentration higher than or equal to the cut-off value was defined as MDS-OAβ positive.
MDS-OAβ positivity could discriminate amyloid PET positivity with the AUC value of 0.855 (95% CI 0.776-0.933). Adding MDS-OAβ positivity to prediction models including age, MMSE score, and APOE ε4 status improved performance up to the AUC value of 0.926 (95% CI 0.871-0.980).
The Aβ oligomerization tendency in plasma could predict amyloid PET positivity with high performance, and, when it is combined with age, MMSE score, and APOE ε4 status, predictability was improved substantially. This suggests the potential of MDS-OAβ as a useful initial stage test in the clinical and research fields of AD.
Summary
Vibrio vulnificus is a halophilic estuarine bacterium that causes fatal septicaemia and necrotizing wound infections. We tested whether V. vulnificus produces signalling molecules ...(autoinducer 1 and/or 2) stimulating Vibrio harveyi quorum‐sensing system 1 and/or 2. Although there was no evidence for signalling system 1, we found that V. vulnificus produced a signalling activity in the culture supernatant that induced luminescence expression in V. harveyi through signalling system 2. Maximal autoinducer 2 (AI‐2) activity was observed during mid‐exponential to early stationary phase and disappeared in the late stationary phase when V. vulnificus was grown in heart infusion broth containing 2.5% NaCl. V. vulnificus showed increased signalling activity when it was cultured in the presence of glucose (0.5%) and at low pH (pH 6.0). From a cosmid library of V. vulnificus type strain ATCC 29307, we have identified the AI‐2 synthase gene (luxSVv) showing 80% identity with that of V. harveyi (luxSVh) at the amino acid level. To investigate the pathogenic role of luxSVv, a deletion mutant of the clinical isolate V. vulnificus MO6‐24/O was constructed. The luxSVv mutant showed a significant delay in protease production and an increase in haemolysin production. The decreased protease and increased haemolysin activities were restored to the isogenic wild‐type level by complementation with the wild‐type luxSVv allele. The change in phenotypes was also complemented by logarithmic phase spent media produced by the wild‐type bacteria. Transcriptional activities of the haemolysin gene (vvhA) and protease gene (vvpE) were also observed in the mutant using chromosomal PvvhA::lacZ and PvvpE::lacZ transcriptional reporter constructs: transcription of vvhA was increased and of vvpE decreased by the mutation. The mutation resulted in an attenuation of lethality to mice. Intraperitoneal LD50 of the luxSVv mutant increased by 10‐ and 750‐fold in ferric ammonium citrate‐non‐overloaded and ferric ammonium citrate‐overloaded mice respectively. The time required for the death of mice was also significantly delayed in the luxSVv mutant. Cytotoxic activity of the organism against HeLa cells, measured by lactate dehydrogenase (LDH) release assay, was also decreased significantly by the mutation. Taken together, the V. vulnificus LuxS quorum‐sensing system seems to play an important role in co‐ordinating the expression of virulence factors.
The sintering behavior of calcium pyrophosphate (CPP, Ca
2P
2O
7)-doped
β-tricalcium phosphate TCP, Ca
3(PO
4)
2, prepared by solid state reaction, was investigated in-situ, using dilatometry. Pure
...β-TCP undergoes phase transition to α-TCP at about 1200°C; hence pure
β-TCP ceramics should be sintered below 1200°C. Pure
β-TCP sintered body can achieve a relative density of only 86% when sintered at 1150°C. However, the addition of CPP in the range of 0.5–3
wt% delays phase transition of
β-TCP and enables sintering of
β-TCP at 1200°C without a phase transformation to
α-TCP. Due to this effect of CPP added to TCP, CPP-doped
β-TCP ceramics with relative density over 95% could be obtained when sintered at 1200°C for 2
h.
The sintering behavior, mechanical properties and biocompatibility of magnesia (MgO)-doped HA/TCP biphasic ceramics were studied. Pure HA/TCP ceramics showed poor sinterability due to the phase ...transformation of
β- to
α-TCP. MgO-doped HA/TCP ceramics showed high density without any phase transformation of
β-TCP up to 1300°C, for MgO dopants incorporated into the
β-TCP preferably and increased thermal stability of
β-TCP. However, the addition of MgO higher than a critical content, suppressed grain growth of HA/TCP ceramics and lowered sinterability. The optimum amount of MgO doping was 1
wt%, which lead 99% relative density and higher mechanical properties than HA or
β-TCP ceramics. From in vitro test and in vivo test, 1
wt% MgO-doped HA/TCP ceramics showed a good biocompatibility without cytotoxicity. After implantation under the muscle of rabbits,
β-TCP phase was dissolved from the surface and a biological apatite covered the surface. These results proved that MgO addition increased drastically the sintering and mechanical properties of HA/
β-TCP ceramics without altering the biological safety and biocompatibility of the original composite.
Diphosphoinositol pentakisphosphate (IP7) is critical for the exocytotic capacity of the pancreatic β-cell, but its regulation by the primary instigator of β-cell exocytosis, glucose, is unknown. The ...high Km for ATP of the IP7-generating enzymes, the inositol hexakisphosphate kinases (IP6K1 and 2) suggests that these enzymes might serve as metabolic sensors in insulin secreting β-cells and act as translators of disrupted metabolism in diabetes. We investigated this hypothesis and now show that glucose stimulation, which increases the ATP/ADP ratio, leads to an early rise in IP7 concentration in β-cells. RNAi mediated knock down of the IP6K1 isoform inhibits both glucose-mediated increase in IP7 and first phase insulin secretion, demonstrating that IP6K1 integrates glucose metabolism and insulin exocytosis. In diabetic mouse islets the deranged ATP/ADP levels under both basal and glucose-stimulated conditions are mirrored in both disrupted IP7 generation and insulin release. Thus the unique metabolic sensing properties of IP6K1 guarantees appropriate concentrations of IP7 and thereby both correct basal insulin secretion and intact first phase insulin release. In addition, our data suggest that a specific cell signaling defect, namely, inappropriate IP7 generation may be an essential convergence point integrating multiple metabolic defects into the commonly observed phenotype in diabetes.
Display omitted
•Glucose increases IP7 levels transiently through IP6K1 in pancreatic β-cells.•IP6K1 decodes glucose-driven increases in ATP/ADP ratio into 1st phase insulin release.•IP7 production and insulin release mirror perturbed metabolism in diabetic islets.•IP6K1 acts as a β-cell metabolic sensor under normal and pathological conditions.
The inositol pyrophosphate, diphosphoinositol pentakisphosphate (IP7), is thought to negatively regulate the critical insulin signaling protein Akt/PKB. Knockdown of the IP7-generating inositol ...hexakisphosphate kinase 1 (IP6K1) results in a concomitant increase in signaling through Akt/PKB in most cell types so far examined. Total in vivo knockout of IP6K1 is associated with a phenotype resistant to high-fat diet, due to enhanced Akt/PKB signaling in classic insulin regulated tissues, counteracting insulin resistance. In contrast, we have shown an important positive role for IP6K1 in insulin exocytosis in the pancreatic β-cell. These cells also possess functional insulin receptors and the feedback loop following insulin secretion is a key aspect of their normal function. Thus we examined the effect of silencing IP6K1 on the activation of Akt/PKB in β-cells. Silencing reduced the glucose-stimulated increase in Akt/PKB phosphorylation on T308 and S473. These effects were reproduced with the selective pan-IP6K inhibitor TNP. The likely explanation for IP7 reduction decreasing rather than increasing Akt/PKB phosphorylation is that IP7 is responsible for generating the insulin signal, which is the main source of Akt/PKB activation. In agreement, insulin receptor activation was compromised in TNP treated cells. To test whether the mechanism of IP7 inhibition of Akt/PKB still exists in β-cells, we treated them at basal glucose with an insulin concentration equivalent to that reached during glucose stimulation. TNP potentiated the Akt/PKB phosphorylation of T308 induced by exogenous insulin. Thus, the IP7 regulation of β-cell Akt/PKB is determined by two opposing forces, direct inhibition of Akt/PKB versus indirect stimulation via secreted insulin. The latter mechanism is dominant, masking the inhibitory effect. Consequently, pharmacological strategies to knock down IP6K activity might not have the same positive output in the β-cell as in other insulin regulated tissues.
Display omitted
•Silencing IP6K1 inhibits glucose-induced Akt/PKB activation in pancreatic β-cells.•A pan IP6K inhibitor, TNP, also reduces glucose-stimulated Akt/PKB phosphorylation.•IP6K1 inhibition impairs Akt/PKB by curtailing insulin secretion and its autocrine feedback.•There are both positive and negative actions of IP7 on Akt/PKB.•The dominant positive action of IP7 on Akt/PKB is due to unique insulin autocrine feedback.
Mitochondria in a variety of cell types respond to physiological Ca
2+ oscillations in the cytosol dynamically with Ca
2+ uptakes. In heart cells, mitochondrial Ca
2+ uptakes occur by a ruthenium ...red-sensitive Ca
2+ uniporter (CaUP), a rapid mode of Ca
2+ uptake (RaM) and a ryanodine receptor (RyR) localized in the inner mitochondrial membrane (IMM). Three subtypes of RyRs have been described and cloned, however, the subtype identity of the mitochondrial ryanodine receptor (mRyR) is unknown. Using subtype specific antibodies, we characterized the mRyR in the IMM from rat heart as RyR1. These results are substantiated by the absence of RyR protein in heart mitochondria from RyR1 knockout mice. The bell-shape Ca
2+-dependent
3Hryanodine binding curve and its modulation by caffeine and adenylylmethylenediphosphonate (AMPPCP) give further evidence that mRyR functions pharmacologically like RyR1. Ryanodine prevents mitochondrial Ca
2+ uptake induced by raising extramitochondrial Ca
2+ to 10 μM. Similarly, ryanodine inhibits oxidative phosphorylation stimulated by 10 μM extramitochondrial Ca
2+. In summary, our results show that the mRyR in cardiac muscle has similar biochemical and pharmacological properties to the RyR1 in the sarcoplasmic reticulum (SR) of skeletal muscle. These results could also suggest an efficient mechanism by which mitochondria sequesters Ca
2+ via mRyR during excitation–contraction coupling to stimulate oxidative phosphorylation for ATP production to meet metabolic demands. Thus, the mRyR functions as a transducer for excitation–metabolism coupling.
PDF
