Development of Al-TiCN nanocomposites via ultrasonic assisted casting route.

This work provides a promising approach to achieve the uniform distribution of TiCN nanoparticles (NPs) in aluminum matrix via a combination of ultrasonic dispersion and fast cooling processing. Microstructure analysis demonstrates that as the cooling rate is increased, the NP distribution in the matrix varies from intergranular to intragranular at micro scale and the NP-matrix interface from incoherent to coherent at nano scale. An analytical model is proposed to unveil the effects of cooling rates on the behavior of NPs at the solidification front. The theoretical analysis reveals that the NP size and cooling rate are the two prominent factors determining the NP distribution during solidification of nanocomposites. The experimental results yield an insight into the understanding of NP-induced microstructural evolution and shed new light on the development of high-performance nanocomposites.

Relatively stable response of fruiting stage to warming and cooling relative to other phenological events.

The timing of the fruit-set stage (i.e., start and end of fruit set) is crucial in a plant's life cycle, but its response to temperature change is still unclear. We investigated the timing of seven phenological events, including fruit-set dates during 3 yr for six alpine plants transplanted to warmer (approximately +3.5°C in soils) and cooler (approximately -3.5°C in soils) locations along an altitudinal gradient in the Tibetan area. We found that fruit-set dates remained relatively stable under both warming and cooling during the 3-yr transplant experiment. Three earlier phenological events (emergence of first leaf, first bud set, and first flowering) and two later phenological events (first leaf coloring and complete leaf coloring) were earlier by 4.8-8.2 d/°C and later by 3.2-7.1 d/°C in response to warming. Conversely, cooling delayed the three earlier events by 3.8-6.9 d/°C and advanced the two later events by 3.2-8.1 d/°C for all plant species. The timing of the first and/or last fruit-set dates, however, did not change significantly compared to earlier and later phenological events. Statistical analyses also showed that the dates of fruit set were not significantly correlated or had lower correlations with changes of soil temperature relative to the earlier and later phenological events. Alpine plants may thus acclimate to changes in temperature for their fruiting function by maintaining relatively stable timings of fruit set compared with other phenological events to maximize the success of seed maturation and dispersal in response to short-term warming or cooling.

MicroRNA-181a-mediated downregulation of AC9 protein decreases intracellular cAMP level and inhibits ATRA-induced APL cell differentiation.

AC9 is one of the adenylate cyclase (AC) isoforms, which catalyze the conversion of ATP to cAMP, an important second messenger. We previously found that the integration of cAMP/PKA pathway with nuclear receptor-mediated signaling was required during all-trans retinoic acid (ATRA)-induced maturation of acute promyelocytic leukemia (APL) cells. Here we showed that AC9 could affect intracellular cAMP level and enhance the trans-activity of retinoic acid receptor. Knockdown of AC9 in APL cell line NB4 could obviously inhibit ATRA-induced differentiation. We also demonstrated that miR-181a could decrease AC9 expression by targeting 3'UTR of AC9 mRNA, finally controlling the production of intracellular cAMP. The expression of miR-181a itself could be inhibited by CEBPα, probably accounting for the differential expression of miR-181a in NB4 and ATRA-resistant NB4-R1 cells. Moreover, we found that AC9 expression was relatively lower in newly diagnosed or relapsed APL patients than in both complete remission and non-leukemia cases, closely correlating with the leukemogenesis of APL. Taken together, our studies revealed for the first time the importance of miR-181a-mediated AC9 downregulation in APL. We also suggested the potential value of AC9 as a biomarker in the clinical diagnosis and treatment of leukemia.

Celecoxib inhibits ß-catenin-dependent survival of the human osteosarcoma MG-63 cell line.

Cyclo-oxygenase (COX)-2 inhibitors may exert antitumour effects through COX-2-independent mechanisms. This study investigated the effects of the COX-2 inhibitor celecoxib on the viability of the human osteosarcoma MG-63 cell line and its ß-catenin signalling pathway. Cell viability and apoptosis were examined in celecoxib-treated cells or after ß-catenin knockdown in vitro. Analyses were performed to detect glycogen synthase kinase (GSK)-3ß, phosphorylated GSK-3ß, ß-catenin, c-Myc and cyclin D1 proteins, and mRNA levels of ß-catenin, c-Myc and CCND1 (encoding cyclin D1). ß-Catenin was shown to be required for MG63 cell survival and celecoxib exerted an inhibitory effect on the viability of cultured MG-63 cells in a time- and dose-dependent manner. ß-Catenin protein decreased in the cytosol and nucleus following celecoxib treatment (from 6 h after initiation of treatment onwards; lowest protein levels were reached at > 72 h). Significant reductions in ß-catenin, c-Myc and CCND1 mRNA were observed. Celecoxib inhibited MG-63 cell viability, possibly by activating GSK-3ß and inhibiting ß-catenin-dependent gene transcription, suggesting a role for celecoxib in osteosarcoma treatment.

Transplanted bone morphogenetic protein/poly(lactic-co-glycolic acid) delayed-release microcysts combined with rat micromorselized bone and collagen for bone tissue engineering.

This study was designed to optimize the preparation of delayed-release microcysts containing bone morphogenetic protein 2 (BMP-2) combined with poly(lactic-co-glycolic acid) (PLGA) and to investigate their osteogenic properties when combined with rat autologous micromorselized bone and collagen. Rat autologous micromorselized bone, collagen and BMP-2/PLGA delayed-release microcysts were implanted in various combinations into the rat gluteus maximus muscle sack model. The following post-operative measurements were made: general observations of the implant site, histological observations, osteogenesis measurements and alkaline phosphatase activity. Autologous micromorselized bone combined with collagen and BMP-2/PLGA delayed-release microcysts demonstrated significantly superior osteogenic properties than any of the other combinations of these three components. These findings suggest that micromorselized bone combined with collagen and BMP-2/PLGA delayed-release microcysts could reduce the quantity of BMP-2 and autologous bone required for these procedures, making their use feasible in human bone restoration.

Dysfunctional mitochondrial respiration in the wobbler mouse brain.

The involvement of mitochondrial dysfunction promoting neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS), has been suggested. Histopathological and biochemical mitochondrial abnormalities have been reported in both sporadic and familial patients and suggest the contention that mitochondria may play a key role promoting ALS. Animal models of ALS provide a unique opportunity to study this incurable and fatal human disease. In the present study we tested the hypothesis that alterations in mitochondrial physiology occur in the brain of wobbler mice. No significant difference was found in the respiratory control index or adenosine diphosphate/oxygen ratio values between isolated mitochondria of wobbler and control mice. When pyruvate and malate were used as substrates, oxygen consumption was decreased significantly by approximately 33% in mitochondria isolated from wobbler mouse brain compared to controls. Oxygen consumption in the presence of ascorbate and N,N,N',N'-tetramethyl-p-phenylenediamine (TMPD) was decreased significantly by approximately 21% in wobbler brain mitochondria compared to controls, which suggests impairment in the function of complex IV. These findings are the first demonstration of mitochondrial respiratory chain dysfunction in the brain of the wobbler mouse.

Cytochrome C is released from mitochondria into the cytosol after cerebral anoxia or ischemia.

Mitochondrial dysfunction may underlie both acute and delayed neuronal cell death resulting from cerebral ischemia. Specifically, postischemic release of mitochondrial constituents such as the pro-apoptotic respiratory chain component cytochrome c could contribute acutely to further mitochondrial dysfunction and to promote delayed neuronal death. Experiments reported here tested the hypothesis that ischemia or severe hypoxia results in release of cytochrome c from mitochondria. Cytochrome c was measured spectrophotometrically from either the cytosolic fraction of cortical brain homogenates after global ischemia plus reperfusion, or from brain slices subjected to severe hypoxia plus reoxygenation. Cytochrome c content in cytosol derived from cerebral cortex was increased after ischemia and reperfusion. In intact hippocampal slices, there was a loss of reducible cytochrome c after hypoxia/ reoxygenation, which is consistent with a decrease of this redox carrier in the mitochondrial pool. These results suggest that cytochrome c is lost to the cytosol after cerebral ischemia in a manner that may contribute to postischemic mitochondrial dysfunction and to delayed neuronal death.

Rapid preconditioning protects rats against ischemic neuronal damage after 3 but not 7 days of reperfusion following global cerebral ischemia.

Earlier studies indicated that sublethal ischemic insults separated by many hours may "precondition" and, thereby, protect tissues from subsequent insults. In Wistar rats, we examined the hypothesis tht ischemic preconditioning (IPC) can improve histopathological outcome even if the "conditioning" and "test" ischemic insults are separated by only 30 min. Normothermic (36.5-37 degrees C) global cerebral ischemia was produced by bilateral carotid artery ligation after lowering mean systemic blood pressure. The conditioning ischemic insult lasted 2 min and was associated with a time sufficient to provoke "anoxic depolarization" (AD) (i.e., the abrupt maximal increase in extracellular potassium ion activity). After 30 min of reperfusion, 10-min test ischemia was produced, and histopathology was assessed 3 and 7 days later. After 3 days of reperfusion, neuroprotection was most robust in the left lateral, middle and medial subsections of the hippocampal CA1 subfield and in the cortex, where protection was 91, 76, 70 and 86%, respectively. IPC also protected the right lateral, middle and medial subsections of the hippocampal CA1 region. These data demonstrate that neuroprotection against acute neuronal injury can be achieved by conditioning insults followed by only short (30 min) periods of reperfusion. However, neuroprotection almost disappeared when reperfusion was continued for 7 days. When test ischemia was decreased to 7 min, a clear trend of neuroprotection by IPC was observed. These data suggest that subsequent rescue of neuronal populations could be achieved with better understanding of the neuroprotective mechanisms involved in this rapid IPC model.

Molecular executors of cell death--differential intrarenal expression of Fas ligand, Fas, granzyme B, and perforin during acute and/or chronic rejection of human renal allografts.

Two distinct cytolytic pathways have been characterized: one in which the interaction between the Fas antigen and its ligand results in apoptosis, and another in which the pore forming protein perforin and the serine protease granzyme B contribute to DNA fragmentation and cell death. We investigated intrarenal expression of these molecular executors of cell death in light of the potential participation of cytolytically active cellular elements in the antiallograft repertory. Reverse transcriptase-polymerase chain reaction was used to identify intrarenal expression of Fas antigen, Fas ligand, granzyme B and perforin in eighty human renal allograft biopsies; mRNA display was correlated with the Banff histological diagnosis of renal allografts. Our studies demonstrate that: (1) intrarenal expression of Fas ligand mRNA and of granzyme B mRNA are correlates of acute but not chronic rejection; (2) Fas ligand mRNA is not detectable in allografts in the absence of rejection; (3) intrarenal coexpression of members of each lytic pathway (Fas ligand and Fas, granzyme B, and perforin) and that of both pathways (e.g., Fas ligand and granzyme B) are correlates of acute rejection; and (4) a direct correlation exists between the histological severity of acute rejection and intrarenal coexpression of mRNA encoding Fas ligand, Fas, granzyme B, and perforin. Our studies identify, for the first time, the differential expression of the two major lytic pathways in acute and chronic allograft rejection and suggest that specific therapy directed at the cytotoxic attack molecules might be efficacious in the prevention and/or treatment of acute rejection.

Intragraft TGF-beta 1 mRNA: a correlate of interstitial fibrosis and chronic allograft nephropathy.

Chronic allograft nephropathy is a relentlessly progressive process and a major cause of long-term graft dysfunction and ultimate failure. Interstitial fibrosis, tubular atrophy, and glomerular and vascular lesions characterize this mechanistically unresolved disorder. Given the prominent role of TGF-beta 1 in tissue repair and in fibrosis, we have explored the hypothesis that fibrosis and chronic allograft nephropathy would be distinguished by intragraft TGF-beta 1 mRNA expression. This postulate was tested by mRNA phenotyping of RNA isolated from 127 human renal allograft biopsies. Reverse transcription assisted polymerase chain reaction was used to amplify and identify ingraft gene expression. Our investigation demonstrated a significant correlation between intragraft TGF-beta 1 mRNA display and renal allograft interstitial fibrosis and chronic allograft nephropathy. In contrast, intragraft expression of mRNA encoding immunoregulatory cytokines, IL-2, IFN-gamma, IL-4, IL-10, or cytotoxic attack molecules, granzyme B and perforin was not a correlate of interstitial fibrosis or chronic allograft nephropathy. Our studies identify, for the first time, a significant association between intragraft TGF-beta 1 mRNA expression and renal allograft interstitial fibrosis, and advance a candidate molecular mechanism for chronic allograft nephropathy.

Intragraft expression of IL-10 messenger RNA: a novel correlate of renal allograft rejection.

A major conceptual advance is the formulation that type I cytokines (such as IL-2 and IFN-gamma) enhance cellular immunity and are host-protective, and that type II cytokines (such as IL-4 and IL-10) dampen cellular immunity and facilitate the progression of infection. We have explored the intragraft expression of type I and type II cytokines during human renal allograft rejection. RNA was isolated from 98 allograft biopsies, and reverse transcription-PCR was used to amplify and identify intragraft expression of mRNA encoding IL-2, IFN-gamma, IL-4, or IL-10. Intragraft expression of IL-7 mRNA and TGF-beta 1 mRNA was also investigated. Our investigation demonstrated that: (a) intragraft expression of IL-10 mRNA and IL-2 mRNA are significant correlates of acute rejection; (b) IL-4, IL-7, IFN-gamma and TGF-beta 1 mRNA expression do not correlate with acute rejection; and (c) IL-10 does not prevent in vivo expression of IFN-gamma, IL-2, IL-7, or TGF-beta 1. Our studies identify, for the first time, a significant association between intragraft IL-10 mRNA expression and acute rejection, and suggest that treatment strategies capable of constraining IL-10 expression might be of value in the prevention of acute rejection.

[Morphine-3-glucuronide (M3G) potentiates noxious stimulus-evoked Fos protein-like immunoreactivity in the rat spinal cord].

The effect of morphine-3-glucuronide (M3G) on noxious stimulus-evoked Fos protein-like immunoreactivity in the rat spinal cord were assessed by ABC method. It was found that a dose-dependent increase of Fos-like immunoreactive neurons could be induced by M3G intrathecal injection followed by formaline injection into hindpaw. With high dosage M3G (1.1 x 10(-7) mole), dense Fos-like labelling was found in the superficial and the deep dorsal horn bilaterally, While with low dosage M3G (5.4 x 10(-8) and 1.1 x 10(-8) mole), most of the positively labelled neurons were only found in laminae I and II of the ipsilateral dorsal horn to the injured paw. The above results revealed that M3G exerts a potentiating effect on the noxious stimulus-evoked Fos protein-like immunoreactivity in the rat spinal cord.

N-nitroso compounds in fresh gastric juice and their relation to intragastric pH and nitrite employing an improved analytical method.

In order to examine further the relationship between intragastric N-nitrosation, gastric pH and nitrite, 457 fresh, fasting gastric juice samples were analysed for total N-nitroso compounds (NOC) and nitrite concentrations using a recently described improved assay method. Nitrite in log values was linearly related to intragastric pH (r = 0.887, P < 0.01) with a regression equation log[nitrite] (mumol/l) = 0.489 x pH - 2.209. Significantly higher NOC concentrations were found at intragastric pH ranges of 1.13-2.99 (mean +/- SE: 1.45 +/- 0.17 mumol/l, P < 0.05) and 6.00-8.42 (3.57 +/- 0.33 mumol/l, P < 0.01) compared with that at pH 3.00-5.99 (1.02 +/- 0.12 mumol/l). NOC concentration was significantly related to log nitrite concentration at both the low pH range 1.13-4.99 (r = 0.169, P < 0.01) and the high pH range 5.00-8.42 (r = 0.450, P < 0.01). The results in the present study confirm that both acid-catalysed N-nitrosation and biologically-catalysed N-nitrosation occur in the human stomach. However, great variations in nitrite and NOC concentrations were observed in both low and high pH samples, indicating that, as expected, both the acid-catalysed N-nitrosation and biologically-catalysed N-nitrosation processes are markedly affected by factors other than intragastric pH and nitrite.

Instability of N-nitroso compounds in gastric juice and preliminary results from analyses of fresh samples by using an improved analytical method.

Using an improved method for determination of total N-nitroso compounds (NOC), we examined the stability of those compounds in fresh gastric juice samples during storage and the effects of the addition of 2% sulphamic acid on NOC concentration in 212 samples. The NOC levels in fresh samples decreased very rapidly at -20 degrees C, especially during the first 24 hours of storage (P < 0.01), and nitrite concentration also showed a decreasing trend during storage. The addition of sulphamic acid significantly reduced NOC levels from 1.97 +/- 0.21 to 1.10 +/- 0.12 mumol/l (mean +/- SE, P < 0.01), especially in samples of initially high pH. However, in some individual samples (16.5%) the NOC levels actually increased by 14.1% (P < 0.01). The results from analysis of NOC in 212 fresh samples in relation to pH demonstrated two significantly higher peaks of NOC concentrations at intragastric pH ranges 1.1 to 2.99 (P < 0.05) and 6.0 to 7.9 (P < 0.01). There was a significant relationship between nitrite level and intragastric pH (r = 0.480, P < 0.01), the nitrite concentration increasing dramatically when the pH exceeded 6.0. The present study suggest that a major proportion of the unidentified NOC formed through intragastric nitrosation is labile NOC; if the true concentration of NOC is to be determined it is therefore essential to analyse fresh gastric juice samples directly after collection and without pretreatment. It will also be necessary to characterize those labile NOC in order to study further the mechanism of endogenous N-nitrosation in man and its relation to human carcinogenesis.

Effects of fruit juices, processed vegetable juice, orange peel and green tea on endogenous formation of N-nitrosoproline in subjects from a high-risk area for gastric cancer in Moping County, China.

The effects of four fruit juices, processed vegetable juice, orange peel, green tea and low dose vitamin C on endogenous N-nitrosation in 86 subjects from a high-risk area for gastric cancer in Moping County, China were studied using urinary excretion of N-nitrosoproline (NPRO) as an indicator. After ingestion of 300 mg L-proline, urinary excretion of NPRO was significantly increased from a baseline of 2.5 +/- 1.6 micrograms/day to 8.7 +/- 6.2 micrograms/day. (P < 0.001). Vitamin C (75 mg) administration significantly reduced NPRO formation (62.3%, P < 0.002) although NPRO excretion remained higher than the baseline level (4.2 +/- 1.3 vs 2.2 +/- 1.2 micrograms/day, P < 0.001). Intake of fruit juices and green tea extracts (containing 75 mg vitamin C) or of orange peel powder (containing 3 mg vitamin C) together with 300 mg L-proline inhibited NPRO formation effectively to the baseline level or to levels significantly lower than the baseline level (P < 0.05-0.005). A processed juice of a number of vegetables (300 ml) significantly catalysed endogenous nitrosation (14.7 +/- 11.8 vs 9.4 +/- 4.7 micrograms/day, P < 0.05). Endogenous N-nitrosation was unaffected by the presence of intragastric lesions. The present study shows that endogenous nitrosation in this population is profoundly affected by environmental factors and that inhibitors, such as vitamin C, alpha-tocopherol and other non-nutritive compounds in the foods do inhibit endogenous nitrosation either synergistically or in an additive manner. The significance of fruits and vegetables in prevention of human cancers is discussed.

Hypothesis on the relationship between gastric cancer and intragastric nitrosation: N-nitrosamines in gastric juice of subjects from a high-risk area for gastric cancer and the inhibition of N-nitrosamine formation by fruit juices.

The concentration of N-nitrosamines (NNA) in gastric juice was determined as an indicator of intragastric N-nitrosation in 85 subjects from a high-risk area for gastric cancer (GC) to examine the relationship between N-nitroso compounds (NOC), pH and intragastric lesions under strictly controlled conditions. Mean gastric pH in subjects with GC or dysplasia (Group GD, 5.0 +/- 2.7) was higher than that from subjects with intestinal metaplasia (Group IM, 3.8 +/- 2.1, p = 0.068) and significantly higher than in those with normal mucosa or superficial gastritis (Group NS, 2.6 +/- 1.9, p < 0.001). No significant difference (p > 0.1) was found in total NNA concentrations between the three groups (GD 1.81 +/- 1.05 micrograms/l, IM 1.46 +/- 0.79 micrograms/l, NS 1.56 +/- 1.38 micrograms/l). However, two obvious peaks of nitrosation were observed at pH ranges of < 2.0 and 5.5-7.5. These observations were confirmed by using the N-nitrosoproline test in the same subjects under the same conditions (r = 0.772, p < 0.05). These results indicate that intragastric nitrosation can occur in both acidic and nearly neutral conditions. The first peak is related to acid-catalysed nitrosation (ACN) and the second is related to biologically catalysed nitrosation (BCN). According to these and other published results the hypothesis that there are two basic mechanisms, ACN and BCN, for intragastric N-nitrosation in humans is explored. Gastric carcinogenesis in high-risk areas is more likely to be related to intragastric NOC formed by ACN, compared to low-risk areas where it is more likely to be related to intragastric NOC formed by BCN. Fruit juices and orange peel significantly inhibited intragastric nitrosation by both ACN and BCN.

[Study on relationship between occurrence of intragastric lesions and drinking water and nitrate intake via water in the inhabitants from a high-risk area for stomach cancer].

According to results of endoscopic and pathological evaluations of gastric mucosa, we investigated some aspects of drinking water for three groups of subjects with various intragastric lesions from a high-risk area (Moping County) for stomach cancer. Their nitrate intakes via drinking water were estimated. The results showed that the occurrence of stomach cancer and other intragastric lesions in these subjects was closely related to their drinking water quality and nitrate intake via water.