Neutropenia and survival outcomes in metastatic colorectal cancer patients treated with trifluridine/tipiracil in the RECOURSE and J003 trials.

BACKGROUND: The phase II J003 (N = 169) and phase III RECOURSE (N = 800) trials demonstrated a significant improvement in survival with trifluridine (FTD)/tipiracil (TPI) versus placebo in patients with refractory metastatic colorectal cancer. This post hoc analysis investigated pharmacokinetic data of FTD/TPI exposure and pharmacodynamic markers, such as chemotherapy-induced neutropenia (CIN) and clinical outcomes. PATIENTS AND METHODS: A total of 210 patients from RECOURSE were enrolled in this substudy. A limited sampling approach was used, with three pharmacokinetic samples drawn on day 12 of cycle 1. Patients were categorized as being above or below the median area under the plasma concentration-time curve (AUC) for FTD and TPI. We conducted a post hoc analysis using the entire RECOURSE population to determine the correlations between CIN and clinical outcome. We then carried out a similar analysis on the J003 trial to validate the results. RESULTS: In the RECOURSE subset, patients in the high FTD AUC group had a significantly increased CIN risk. Analyses of the entire population demonstrated that FTD/TPI-treated patients with CIN of any grade in cycles 1 and 2 had significantly longer median overall survival (OS) and progression-free survival (PFS) than patients who did not develop CIN and patients in the placebo group. Patients who required an FTD/TPI treatment delay had increased OS and PFS versus those in the placebo group and those who did not develop CIN. Similar results were obtained in the J003 cohort. CONCLUSIONS: In RECOURSE, patients with higher FTD drug exposure had an increased CIN risk. FTD/TPI-treated patients who developed CIN had improved OS and PFS versus those in the placebo group and those who did not develop CIN. Similar findings were reported in the J003 cohort, thus validating the RECOURSE results. The occurrence of CIN may be a useful predictor of treatment outcomes for FTD/TPI-treated patients. CLINICALTRIALS. GOV IDENTIFIER: NCT01607957 (RECOURSE). JAPAN PHARMACEUTICAL INFORMATION CENTER NUMBER: JapicCTI-090880 (J003).

Abnormal asymmetries in subcortical brain volume in schizophrenia.

Subcortical structures, which include the basal ganglia and parts of the limbic system, have key roles in learning, motor control and emotion, but also contribute to higher-order executive functions. Prior studies have reported volumetric alterations in subcortical regions in schizophrenia. Reported results have sometimes been heterogeneous, and few large-scale investigations have been conducted. Moreover, few large-scale studies have assessed asymmetries of subcortical volumes in schizophrenia. Here, as a work completely independent of a study performed by the ENIGMA consortium, we conducted a large-scale multisite study of subcortical volumetric differences between patients with schizophrenia and controls. We also explored the laterality of subcortical regions to identify characteristic similarities and differences between them. T1-weighted images from 1680 healthy individuals and 884 patients with schizophrenia, obtained with 15 imaging protocols at 11 sites, were processed with FreeSurfer. Group differences were calculated for each protocol and meta-analyzed. Compared with controls, patients with schizophrenia demonstrated smaller bilateral hippocampus, amygdala, thalamus and accumbens volumes as well as intracranial volume, but larger bilateral caudate, putamen, pallidum and lateral ventricle volumes. We replicated the rank order of effect sizes for subcortical volumetric changes in schizophrenia reported by the ENIGMA consortium. Further, we revealed leftward asymmetry for thalamus, lateral ventricle, caudate and putamen volumes, and rightward asymmetry for amygdala and hippocampal volumes in both controls and patients with schizophrenia. Also, we demonstrated a schizophrenia-specific leftward asymmetry for pallidum volume. These findings suggest the possibility of aberrant laterality in neural pathways and connectivity patterns related to the pallidum in schizophrenia.

Effect of active packaging on low-sodium restructured chicken steaks.

Contributing for a healthier lifestyle, the technology of active and biodegradable packaging with antimicrobial and/or antioxidants compounds and reduced sodium intake have been increasingly applied in meat and meat products. Thus, the objective of this research was to assess the effectiveness of oregano essential oil (OEO) and potassium sorbate incorporated in packaging applied to the restructured chicken steaks with 40 % reduction in sodium chloride in frozen storage for 150 days. The composition of packaging did not influence moisture, crude protein, total lipids, ash, sodium and potassium content as well as pH evaluated on days 0 and 150. Salty taste was the only significant indication in the sensory analysis (p < 0.05). The use of 1 % and 0.5 % OEO incorporated in packaging reduced rancidity through lipid oxidation and can be regarded as an active antioxidant; the use of oregano or potassium sorbate in active films caused the development delay effect E. coli. Thus, the use of active packaging may maintain the product quality.

Chair rising exercise is more effective than one-leg standing exercise in improving dynamic body balance: a randomized controlled trial.

A randomized controlled trial was conducted to compare the effect of a one-leg standing exercise and a chair-rising exercise on body balance in patients with locomotive disorders. Thirty ambulatory patients (mean age: 66.6 years) were randomly divided into two groups (n=15 in each group): a one-leg standing exercise group and a chair-rising exercise group. All the participants performed calisthenics of the major muscles, a tandem gait exercise, and a stepping exercise. The exercises were performed 3 days per week, and the study period was 5 months. Physical function was evaluated at baseline and at one-month intervals. No significant differences in the baseline characteristics were observed between the two groups. After the 5-month exercise program, the timed up and go, one-leg standing time, and tandem gait time improved significantly in the one-leg standing exercise group, while the walking time and chair-rising time in addition to above parameters improved significantly in the chair-rising exercise group. The improvements in the walking time, chair-rising time, and tandem gait time were significantly greater in the chair-rising exercise group than in the one-leg standing exercise group. The present study showed that the chair-rising exercise was more effective than the one-leg standing exercise for improving walking velocity and dynamic body balance.

Enhancement of gene expression by transcriptional activation using doxorubicin-loaded liposome/pDNA complexes.

Gene therapy is a promising treatment option for cancers generated by mutation of oncogenes or tumor suppressor genes. The transcriptional process is activated by doxorubicin (DXR), and gene expression efficiency followed by gene transfection can be enhanced by the combination-use of DXR. Therefore, co-encapsulation of plasmid DNA (pDNA) and DXR into non-viral gene carriers can enhance gene expression. Here, we prepared DXR-loaded liposome/pDNA complexes (DXR-loaded PEGylated lipoplexes) by co-encapsulating pDNA and DXR into liposomes. Gene expression was enhanced by DXR encapsulation into lipoplexes in colon-26 cells and cultured mouse macrophages, and this gene expression level was significantly higher than that obtained by the combination of PEGylated lipoplexes and free DXR. Moreover, the activation profiles of transcriptional factors induced by DXR-loaded lipoplexes were different from those induced by free DXR; therefore, co-encapsulation of pDNA and DXR into gene carriers might be contributed to effective enhancement of gene expression. These findings provide a new approach for achieving effective gene transfection using PEGylated lipoplexes.

Position of the mandibular canal before and after bilateral sagittal split ramus osteotomy: a cone beam computed tomographic study.

The aim of this study was to evaluate the position of the mandibular canal (MC) before and after bilateral sagittal split ramus osteotomy (BSSRO) using cone-beam computed tomography (CT), and to compare the position of the MC in Class II and Class III patients in the preoperative period. Patients were divided into two groups: Class II (n = 38) and Class III (n = 41). Measurements of the superior, inferior, buccal, and lingual distances of the MC in relation to the cortical bone were taken at three levels in the proximal segment of the mandible. Results were analysed using the Kruskal-Wallis test (p < 0.05). In the Class II group the superior distance of the MC at levels 2 and 3, and the inferior distance at level 3 significantly decreased after BSSRO. In the Class III group, no significant differences were found at any level, and the inferior distances at all levels were smaller preoperatively than those in the Class II group. In the Class II group the position of the MC altered in relation to superior and inferior cortical bone after BSSRO. However, the position of the MC remained stable in the Class III group. Our results also suggest a deeper cut in inferior cortical bone in Class III patients.

Purification and characterization of a human membrane protein that activates the alternative complement pathway and allows the deposition of homologous complement C3.

A human myeloid cell subline, P39+, is found to be a target for human complement (C) via the alternative pathway and to allow the deposition of multiple C3 fragments on its membranes, though expressing the complement regulatory proteins decay-accelerating factor and membrane cofactor protein. The parent cell line, P39-, which is phenotypically similar to the P39+ subline, does not allow the deposition of homologous C3 fragments. In this study, we established a monoclonal antibody, M161 Ab, which reacted with P39+ but not P39- cells. This Ab recognized a 43-kD protein in P39+ cell lysate transblotted onto nitrocellulose. Using this Ab as a probe, we purified the 43-kD protein, namely, M161 antigen (Ag). M161 Ag had a basic isoelectric point (pI), 9.3-9.4 by chromatofocusing, and was precipitated as an insoluble material at the pI point. The purified M161 Ag was a single-chain protein and did not possess N- or O-linked carbohydrates. When the purified M161 Ag was transblotted onto nitrocellulose and incubated with Mg(2+)-EGTA serum, human C3 fragments were efficiently deposited on M161 Ag. The major species of the deposited C3 fragments was C3b. Furthermore, the C3 fragments bound to the M161 Ag were detached by 1 M hydroxylamine, suggesting that a covalent ester linkage sustains M161 Ag-C3b interaction. NH2-terminal amino acid analysis revealed that M161Ag is a novel membrane protein. Hence, it appeared that M161 Ag is a potent activator of human alternative complement pathway on human cells that activates homologous C3 and allows the deposition of C3b on itself. Thus, under some conditions, homeostasis of complement is maintained even on human cells, not only by the complement regulatory proteins, but also by membrane C3-activating molecules on which C3b is deposited.

Asymmetrical interhemispheric fiber tracts in patients with hemimegalencephaly on diffusion tensor magnetic resonance imaging.

OBJECTIVE: The internal structures of cerebral white matter in patients with hemimegalencephaly have not yet been investigated except for one, which evaluated aberrant fibers. We examined interhemispheric fiber tracts (FT) passing through the corpus callosum using magnetic resonance (MR) diffusion tensor imaging (DTI). METHODS: MR studies, including DTI, were performed in nine consecutive patients with hemimegalencephaly and in 11 patients with West syndrome as disease controls. The interhemispheric FT passing through the corpus callosum were evaluated in six regional geometric subdivisions in every hemimegalencephaly and West syndrome patient (54 and 66 subregions, respectively), and the distribution and volume differences between affected and unaffected hemispheres were all compared. RESULTS: In patients with hemimegalencephaly, interhemispheric FT were symmetrically distributed in 27 (50%) of the 54 corpus callosum subregions. However, the FT were distributed to different areas in the same lobes in 22 (40%) subregions, and to different lobes in five (9%) subregions. FT volumes were symmetrical in 35 (65%) subregions, while FT volumes on the affected side were greater, but less than those on the unaffected side, in 14 (26%) and five (9%) subregions, respectively. In contrast, in the West syndrome patients, interhemispheric FT showed symmetrical distributions and volumes in all regions. CONCLUSION: Asymmetrical interhemispheric FT are often observed in patients with hemimegalencephaly, and DTI was a useful means of elucidating the internal structures of white matter.

Effects of prenatal ethanol exposure on regulation of basal hypothalamic-pituitary-adrenal activity and hippocampal 5-HT1A receptor mRNA levels in female rats across the estrous cycle.

Prenatal ethanol exposure, like other early adverse experiences, is known to alter hypothalamic-pituitary-adrenal (HPA) activity in adulthood. The present study examined the modulatory effects of the gonadal hormones on basal HPA regulation and serotonin Type 1A receptor (5-HT(1A)) mRNA levels in adult female rats prenatally exposed to ethanol (E) compared to that in females from pair-fed (PF) and ad libitum-fed control (C) conditions. We demonstrate, for the first time, long-lasting consequences of prenatal ethanol exposure for basal corticosterone (CORT) regulation and basal levels of hippocampal mineralocorticoid (MR), glucocorticoid (GR) and serotonin Type 1A (5-HT(1A)) receptor mRNA, as a function of estrous cycle stage: (1) basal CORT levels were higher in E compared to C females in proestrus but lower in E and PF compared to C females in estrus; (2) there were no differences among groups in basal levels of adrenocorticotropin (ACTH), estradiol or progesterone; (3) hippocampal MR mRNA levels were decreased in E compared to PF and C females across the estrus cycle, with the greatest effects in proestrus, whereas E (but not PF or C) females had higher hippocampal GR mRNA levels in proestrus than in estrous and diestrus; (4) 5-HT(1A) mRNA levels were increased in E compared to PF and C females in diestrus. That alterations were revealed as a function of estrous cycle stage suggests a role for the ovarian steroids in mediating the adverse effects of ethanol. Furthermore, it appears that ethanol-induced nutritional effects may play a role in mediating at least some of the effects observed. The resetting of HPA activity by early environmental events could be one mechanism linking early life experiences with long-term health consequences. Thus, changes in basal CORT levels, a shift in the MR/GR balance and alterations in 5-HT(1A) receptor mRNA could have important clinical implications for understanding the secondary disabilities, such as an increased incidence of depression, in children with FASD.

Hemimegalencephaly: a study of abnormalities occurring outside the involved hemisphere.

BACKGROUND AND PURPOSE: Hemimegalencephaly is a rare but well-known congenital malformation with ipsilateral enlargement of the hemicerebrum. However, very little is known about changes in structures outside the involved hemisphere in patients with this condition. We investigated morphologic abnormalities occurring outside the affected hemisphere by MR imaging in a large series of patients with hemimegalencephaly. MATERIALS AND METHODS: MR imaging findings for 30 patients with hemimegalencephaly were retrospectively reviewed and evaluated for structures outside the involved hemisphere on routine MR images, such as cranial nerves (I, II, V), brain vessels, subdural and subarachnoid spaces, brain stem, and cerebellum, on both the ipsilateral and contralateral sides. RESULTS: The ipsilateral olfactory and optic nerves were enlarged in 8 (26.7%) and 1 (3.3%) of the 30 patients, respectively, without enlargement on the contralateral side. No asymmetry was noted in the trigeminal nerves. Asymmetric vascular dilations in the ipsilateral cerebral hemisphere were observed in 12 of the 30 patients (40%), in deep cerebral vessels in 11 patients (36.7%), and in superficial cerebral vessels in 8 patients (26.7%). Ipsilateral brain stem and hemicerebellar asymmetric enlargement was detected in 2 patients (6.7%) and 14 patients (46.7%), respectively. Abnormal cerebellar folia were observed on the ipsilateral side in 6 patients (20%) and on the contralateral side in 3 patients (10%). CONCLUSION: Ipsilateral olfactory nerve enlargement, cerebral vascular dilations, cerebellar enlargement, and bilateral or ipsilateral abnormal architecture of the cerebellar folia are often associated with hemimegalencephaly.

Sepiolite as a promising nanoclay for nano-biocomposites based on starch and biodegradable polyester.

The effects of sepiolite addition (0, 1, 3 and 5wt%) were evaluated on dynamic-mechanical behaviour, water uptake, thermal and optical properties of thermoplastic starch (TPS)/poly (butylene adipate-co-terephthalate) (PBAT) nano-biocomposites, with different TPS/PBAT (w/w) ratios and nanofiller contents. The results highlighted the improvement of the dynamic-mechanical behaviour with the addition of sepiolite, producing high performance materials. An increase of 25°C in the Tg of TPS was recorded by DMTA analysis at sepiolite content of 5wt%. The sepiolite influenced the crystallisation of nano-biocomposites, without causing interference in the crystal organisation, evidenced by DSC analysis. The addition of sepiolite nanoclay decreased the water adsorption rate and water adsorption capacity of the corresponding nano-biocomposites. For such multiphase systems, the successful use of natural sepiolite brought a clear benefit, without the need of any modifications or additional processes to produce advanced nano-biocomposites.

Preoperative Cerebral Oxygen Extraction Fraction Imaging Generated from 7T MR Quantitative Susceptibility Mapping Predicts Development of Cerebral Hyperperfusion following Carotid Endarterectomy.

BACKGROUND AND PURPOSE: Preoperative hemodynamic impairment in the affected cerebral hemisphere is associated with the development of cerebral hyperperfusion following carotid endarterectomy. Cerebral oxygen extraction fraction images generated from 7T MR quantitative susceptibility mapping correlate with oxygen extraction fraction images on positron-emission tomography. The present study aimed to determine whether preoperative oxygen extraction fraction imaging generated from 7T MR quantitative susceptibility mapping could identify patients at risk for cerebral hyperperfusion following carotid endarterectomy. MATERIALS AND METHODS: Seventy-seven patients with unilateral internal carotid artery stenosis (≥70%) underwent preoperative 3D T2*-weighted imaging using a multiple dipole-inversion algorithm with a 7T MR imager. Quantitative susceptibility mapping images were then obtained, and oxygen extraction fraction maps were generated. Quantitative brain perfusion single-photon emission CT was also performed before and immediately after carotid endarterectomy. ROIs were automatically placed in the bilateral middle cerebral artery territories in all images using a 3D stereotactic ROI template, and affected-to-contralateral ratios in the ROIs were calculated on quantitative susceptibility mapping-oxygen extraction fraction images. RESULTS: Ten patients (13%) showed post-carotid endarterectomy hyperperfusion (cerebral blood flow increases of ≥100% compared with preoperative values in the ROIs on brain perfusion SPECT). Multivariate analysis showed that a high quantitative susceptibility mapping-oxygen extraction fraction ratio was significantly associated with the development of post-carotid endarterectomy hyperperfusion (95% confidence interval, 33.5-249.7; P = .002). Sensitivity, specificity, and positive- and negative-predictive values of the quantitative susceptibility mapping-oxygen extraction fraction ratio for the prediction of the development of post-carotid endarterectomy hyperperfusion were 90%, 84%, 45%, and 98%, respectively. CONCLUSIONS: Preoperative oxygen extraction fraction imaging generated from 7T MR quantitative susceptibility mapping identifies patients at risk for cerebral hyperperfusion following carotid endarterectomy.

Prenatal ethanol exposure alters the effects of gonadectomy on hypothalamic-pituitary-adrenal activity in male rats.

Prenatal ethanol exposure has marked effects on development of the hypothalamic-pituitary-adrenal (HPA) and -gonadal (HPG) axes. In adulthood, ethanol-treated rats show altered gonadal hormone responses and reproductive function, and increased HPA responsiveness to stressors. Importantly, prenatal ethanol differentially alters stress responsiveness in adult males and females, raising the possibility that the gonadal hormones play a role in mediating prenatal ethanol effects on HPA function. To examine a possible testicular influence on HPA activity in males, we compared the effects of gonadectomy on HPA stress responses of adult male offspring from ethanol, pair-fed (PF) and ad libitum-fed control dams. Intact ethanol-treated rats showed increased adrenocorticotrophic hormone (ACTH) but blunted testosterone and luteinising hormone (LH) responses to restraint stress, and no stress-induced elevation in arginine vasopressin (AVP) mRNA levels compared to those observed in PF and/or control rats. Gonadectomy: (i) significantly increased ACTH responses to stress in control but not ethanol-treated and PF males; (ii) eliminated differences among groups in plasma ACTH and AVP mRNA levels; and (iii) altered LH and gonadotrophin-releasing hormone responses in ethanol-treated males. Taken together, these findings suggest that central regulation of both the HPA and HPG axes are altered by prenatal ethanol exposure, with normal testicular influences on HPA function markedly reduced in ethanol-treated animals. A decreased sensitivity to inhibitory effects of androgens could contribute to the HPA hyperresponsiveness typically observed in ethanol-treated males.

Physicochemical and pharmacokinetic characteristics of cationic liposomes.

After intravenous administration of plasmid DNA (pDNA)/cationic liposome complexes, the gene expression is predominantly observed in the lung due to the physicochemical properties of the liposome complexes and the physiology of the lung. To determine the physicochemical properties and distribution behavior of cationic liposomes for lung-selective drug and/or gene delivery systems, N-[1-(2,3-dioleyloxy)propyl]-n,n,n-trimethylammonium chloride (DOTMA)/cholesterol and 1,2-dioleoyl-3-trimethyl-ammoniopropane (DOTAP)/cholesterol liposomes were studied. The particle sizes of DOTMA/cholesterol and DOTAP/cholesterol liposomes were very similar: 126 and 128 nm, respectively. Furthermore, the zeta potentials of these two liposomes were 51 and 66 mV, respectively. After intravenous injection into mice, both cationic liposomes were rapidly eliminated from the blood circulation and preferentially recovered in the lung. Interestingly, the highest lung accumulation was observed at 1 min, and then, decreased gradually. The distribution characteristics of DOTMA/cholesterol and DOTAP/cholesterol liposomes were almost identical due to the similarities in their physicochemical properties. These results demonstrated that DOTMA/cholesterol and DOTAP/cholesterol liposomes, which possess a positive charge, are promising carriers for lung-selective drug and/or gene delivery systems.

Suppression of TNFalpha production in LPS induced liver failure in mice after intravenous injection of cationic liposomes/NFkappaB decoy complex.

NFkappaB decoy, double stranded oligonucleotides containing NFkappaB binding sequences, inhibits NFkappaB-mediated production of inflammatory cytokines, and therefore NFkappaB decoy has been applied to several diseases. However, naked NFkappaB decoy, which is quickly cleared from the circulation in mice after intravenous injection, is readily absorbed into the systemic circulation. In order to deliver enough NFkappaB decoy for a therapeutic effect, it is necessary to develop a carrier, which enables much more NFkappaKB decoy to transfer to the target cells. In this study, using N-[1-(2,3-dioleyloxy)propyl]-n,n,n-trimethylammonium chloride (DOTMA)/cholesterol (1 :1) liposomes, the therapeutic effect of NFkappaB decoy was investigated in an LPS induced acute hepatitis model mice. The mean diameter of the cationic liposomes/NFkappaB decoy complex was about 70.9 nm and the zeta potential of complex was about 37.4 mV. Tissue distribution was determined by measuring the radioactivity of a cationic liposomes/ [32P] NFkappaB decoy complex after intravenous injection. The cationic liposomes/[32P] NFkappaB decoy complex was rapidly accumulated in the lung and gradually moved to the liver. The therapeutic effect was determined by the serum concentration of TNFalpha in LPS treated mice. The production of TNFalpha was significantly inhibited by cationic liposomes/NFkappaB decoy complex but not by cationic liposomes/random decoy complex or naked NFkappaB decoy. These results suggested that NFkappaB decoy therapy could be achieved using cationic liposomes. This information is of great value for the design of NFkappaB decoy carrier systems.

Intratumoral pharmacokinetics and in vivo gene expression of naked plasmid DNA and its cationic liposome complexes after direct gene transfer.

The pharmacokinetic properties and gene expression of naked plasmid DNA and its cationic liposome complexes were studied after direct intratumoral injection. Using a Walker 256 tissue-isolated tumor perfusion system, we quantified the recovery of naked plasmid DNA and cationic liposome complexes in the tumor, leakage from the tumor surface, and the venous outflow after intratumoral injection. Approximately 50% of naked plasmid DNA had been eliminated from the tumor 2 h after injection, whereas more than 90% of plasmid DNA was retained in the tumor when it was complexed with cationic liposomes. However, the distribution of these complexes in the tumor was restricted to the tissue surrounding the injection site. Pharmacokinetic analysis of the venous outflow profiles suggested that the rate-limiting process that determines the retention of plasmid DNA in the tumor is transferred from the injection site in the tumor tissue and that complexation with cationic liposomes may retard this process. Furthermore, we examined the gene expression of chloramphenicol acetyltransferase DNA constructs (naked pCMV-CAT) and the corresponding cationic liposome [3-beta-(N-(N',N'-dimethylaminoethane)carbamoyl)cholesterol] complexes. A similar level of gene expression was observed in vivo after direct intratumoral injection of naked DNA and its cationic liposome complexes. In both cases, a great variation was observed between tumors, and localization of gene-transduced cells in the tumor tissue was limited to the area in the vicinity of the injection site. Thus, these pharmacokinetic and gene expression studies have demonstrated that cationic liposomes can enhance the retention of injected DNA in the tumor model, whereas cationic liposome complex does not necessarily improve gene expression because of its poor dissemination in this tumor. The present study also suggested that there is a need to control the behavior of the injected naked plasmid DNA and its cationic liposome complexes to ensure better distribution throughout the tumor.

Biodistribution characteristics of mannosylated, fucosylated, and galactosylated liposomes in mice.

The in vivo disposition behavior and pharmacokinetic characteristics of galactosylated (Gal), mannosylated (Man) and fucosylated (Fuc) liposomes were compared in this study. For the preparation of the glycosylated liposomes, cholesten-5-yloxy-N-(4-((1-imino-2-beta-D-thiogalactosyle thyl)amino)a lkyl)formamide (Gal-C4-Chol) (Kawakami et al., Biochem. Biophys. Res. Commun. 252 (1998) 78-83) and its mannosylated and fucosylated derivatives (Man-C4-Chol and Fuc-C4-Chol, respectively) were synthesized. The glycosylated liposomes are composed of distearoylphosphatidylcholine (DSPC), cholesterol (Chol), and Gal-C4-Chol (or Man-C4-Chol or Fuc-C4-Chol) with the molar ratio of 60:35:5. After intravenous injection in mice, these three types of [(3)H]cholesteryl hexadecyl ether-labeled glycosylated liposomes were rapidly eliminated from the circulating blood and preferentially recovered in the liver. In contrast, DSPC/Chol (60:40) liposomes without glycosylation were retained for a long time in the circulating blood. The uptake ratios by parenchymal cells (PC) and nonparenchymal cells (NPC) (PC/NPC ratios) for 0.5% Gal, Man and Fuc liposomes were found to be 15.1, 0.6 and 0.2, respectively. The effect of predosing glycosylated proteins and liposomes on the hepatic uptake of 0.5% (3)H-labeled Gal, Man, and Fuc liposomes was investigated and the results support the conclusion that Gal, Man, and Fuc liposomes are taken up by the liver via asialoglycoprotein receptors in PC, mannose receptors in NPC, and fucose receptors in NPC, respectively. Interestingly, Gal liposomes were taken up by NPC rather than by PC at a high dose (5%). Together with the finding that 5% Gal liposomes inhibit the hepatic uptake of (3)H-labeled Fuc liposomes, this suggests that Gal-liposomes administered at a high dose will also be taken up by fucose receptors in NPC, that are considered to act as galactose particle receptors.

Serum mannan binding protein inhibits mannosylated liposome-mediated transfection to macrophages.

The effects of serum mannan binding proteins (MBP) in the transfection of plasmid DNA/Man-liposome complex via mannose receptor-mediated endocytosis was studied in vitro using cultured mouse peritoneal macrophages. Plasmid DNA encoding luciferase gene was complexed with cationic mannosylated liposomes (Man-liposomes), composed of cholesten-5-yloxy-N-(4-((1-imino-2-D-thiomannosylethyl)amino)alkyl)formamide (Man-C4-Chol) and dioleoyl phosphatidylethanolamine (DOPE). The transfection efficiency, as well as the binding and uptake of the plasmid DNA/Man-liposome complex, was investigated with or without serum MBP. The in vitro transfection efficiency of the complex was significantly reduced on increasing the amount of serum MBP. In addition, the cellular association of the complex was also reduced. These results indicate that serum MBP specifically binds to the mannose moieties on the complex and suppresses its cellular uptake, resulting in inhibition of the gene transfection in macrophages. Such an interaction is an obstacle to mannose receptor-mediated in vivo gene transfer to mannose receptor-positive cells using mannosylated gene carriers.

Involvement of serum mannan binding proteins and mannose receptors in uptake of mannosylated liposomes by macrophages.

The roles of serum mannan binding protein (MBP) and the mannose receptor in the cellular uptake of mannosylated liposomes (Man-liposomes) by macrophages were studied. Man-liposomes were prepared by incorporating cholesten-5-yloxy-N-(4-((1-imino-2-beta-D-thiomannosylethyl)amino)butyl)formamide (Man-C4-Chol) into small unilamellar long circulating liposomes consisting of cholesterol (Chol) and distearoyl phosphatidylcholine (DSPC). In the in vitro cellular uptake study with cultured mouse peritoneal macrophages, [(3)H]Man-liposomes were taken up to a great extent, whereas no significant uptake was observed for [(3)H]cholesterol and DSPC liposomes without Man-C4-Chol (Bare-liposomes). The uptake of [(3)H]Man-liposomes was dose- and temperature-dependent and inhibited by an excess of mannosylated bovine serum albumin, suggesting their specific uptake via membrane mannose receptor-mediated endocytosis. Furthermore, it was demonstrated that (111)In-MBP binds strongly to Man-liposomes based on the recognition of Man-C4-Chol and markedly enhanced their uptake by macrophages. These results are supported by confocal laser microscopic images. In addition, in vivo hepatic uptake of (111)In-MBP was enhanced by Man-liposomes. On the other hand, the uptake of Man-liposomes was significantly reduced by preincubation with serum and further with MBP-depleted serum suggesting inhibitory effects of serum proteins such as albumin on mannose receptor-mediated endocytosis. The involvement of serum-type MBP and membrane mannose receptors in the uptake of Man-liposomes is thus suggested.

Elaboration, morphology and properties of starch/polyester nano-biocomposites based on sepiolite clay.

The incorporation of nano-sized sepiolite clays into thermoplastic starch/poly(butylene adipate-co-terephthalate) (TPS/PBAT) blends has been investigated with the goal of improving the matrix properties. TPS/PBAT nano-biocomposites were elaborated with two different proportions of the polymeric phases. The influence of the sepiolite nanoclays on the mechanical, thermal and structural properties of the corresponding blends was evaluated. SEM images confirmed the good dispersion of the sepiolite clay, with a low occurrence of small aggregates in the polymeric matrix. Wide-angle X-ray diffraction showed no significant alteration of the crystalline structures of PBAT and starch induced by the sepiolite clay. The addition of sepiolite slightly affected the thermal degradation of the nano-biocomposites; however, the mechanical tests revealed an increase in some mechanical properties, demonstrating that sepiolite is a promising nanofiller for TPS-based materials.