Association between Preexisting Sarcopenia and Stroke in Patients with Type 2 Diabetes Mellitus.

OBJECTIVES: This propensity score-matched population-based cohort study compared stroke risk between patients with type 2 diabetes mellitus with and without preexisting sarcopenia. RESEARCH DESIGN AND METHODS: We used data from Taiwan's National Health Insurance Research Database for the period from January 2008 to December 2019. We recruited patients with type 2 diabetes mellitus and categorized them into two groups at a ratio of 1:1 on the basis of diagnosed sarcopenia. The matching variables were age, sex, income level, urbanization level, diabetes severity (adapted Diabetes Complications Severity Index [aDCSI Scores]), Charlson Comorbidity Index (CCI), other comorbidities associated with stroke, smoking status, medication use, and types of antidiabetic medications. The matching process yielded a final cohort of 104,120 patients (52,060 and 52,060 in the sarcopenia and nonsarcopenia groups, respectively) who were eligible for inclusion in subsequent analyses. RESULTS: In the multivariate Cox regression analysis, the adjusted hazard ratio (aHR; 95% CI) of stroke for the sarcopenia diabetes group compared with the control group was 1.13 (1.10, 1.16; P < 0.001), after controlling for age, sex, CCI, and aDCSI scores. The incidence rates of stroke for the sarcopenia and nonsarcopenia groups were 295.98 and 260.68 per 10,000 person-years, respectively. The significant IRR (95% CI) of stroke was 1.14 (1.09, 1.17) for the sarcopenia diabetes group compared with the nonsarcopenic diabetes group. CONCLUSION: Preexisting sarcopenia increased the risk of stroke in patients with type 2 diabetes mellitus.

Deformation Potential and Tensile Strength of Tablets of a Dry Granulated Formulation.

The increase in solid fraction (SF) of a packed granule bed with pressure applied during the in-die compression process results in an evolution of the tablet's matrix and mechanical strength. In this case study, the tensile strength (TS) of a dry granulated microcrystalline cellulose (MCC)/mannitol (MNT)-based formulation was modeled in light of the deformation potential, ∆ (tablet SF - initial granule bed SF). Results showed that the TS of tablets linearly decreased as SF of granules (produced as mini-tablets of an ibuprofen formulation) increased. The formulated granules achieved a measurable tablet strength at a slightly lower critical deformation potential (∆c) than the pure MCC granules. Beyond ∆c, tablet TS increased almost linearly as the deformation potential increased, and the rate was higher for tablets with higher SF. Compared to the simple MCC system, the granules of the MCC/MNT-based formulation were weaker, and TS of tablets increased with deformation potential at a lower rate.

Sirt7 protects chondrocytes degeneration in osteoarthritis via autophagy activation.

OBJECTIVE: Osteoarthritis (OA) is associated with decreased autophagy activity and imbalance of cell homeostasis in chondrocytes (CHs). Sirtuin 7 (Sirt7) is claimed to have the ability to activate the autophagy response. The aim of this study was to explore the function of Sirt7 in the development of OA involving autophagy by culturing human chondrocytes (CHs). PATIENTS AND METHODS: We collected knee joint cartilage from patients undergoing traumatic amputation and arthroscopic knee replacement. Protein and mRNA levels of collagen II, Sirt7, ULK1, Lc3, and Beclin1 were analyzed by Western blot and RT-PCR. CHs were isolated from the traumatic cartilage as a control group, and IL-1ß was used to induce CHs degeneration. The expression of Sirt7 gene was silenced by siRNA and upregulated by recombinant human Sirt7 protein (rh-Sirt7). An autophagy activator Tat-beclin 1 (Tat) was used to activate autophagy in cultural CHs. Expression of collagen II, Sirt7, ULK1, Lc3, and Beclin1 was determined by immunofluorescence, Western blot, and RT-PCR, respectively. RESULTS: The protein and mRNA levels of Collagen II, Sirt7, ULK1, Lc3-II, and Beclin1 expressions were decreased in OA cartilage compared with those in healthy cartilage. IL-1ß degenerated the CHs resulting in a reduction of collagen II, which also downregulated Sirt7, ULK1, Lc3-II, and Beclin1. Sirt7 deficiency accelerated the catabolism of collagen II and weakened the expression of ULK1, Lc3-II, and Beclin1. On the contrary, exogenous rh-Sirt7 played a positive role in these gene expressions. Finally, Tat alleviated the CHs degeneration by upregulating collagen II and activating ULK1, Lc3-II, and Beclin1, but incapable to mediate Sirt7 expression. CONCLUSIONS: Overall, these findings suggested that Sirt7 was suppressed in the degenerated cartilage. Sirt7 deficiency does harm to the autophagy level, affecting CHs metabolism, while the upregulation of Sirt7 activated autophagy and protected CHs degeneration. An appropriate increase in autophagy can protect CHs but has no effect on Sirt7 expression.

IL-9 exacerbates the development of chronic obstructive pulmonary disease through oxidative stress.

OBJECTIVE: To investigate the role of IL-9 in chronic obstructive pulmonary disease (COPD), and to explore its potential mechanism. MATERIALS AND METHODS: A mouse COPD model was established by exposure to cigarette smoke. COPD mice were then randomly assigned into two groups, including: the PBS group and the IL-9 antibody group. The above two groups were treated with phosphate-buffered saline (PBS) or IL-9 injection, respectively. The histopathological changes in lung tissues of mice were observed by hematoxylin-eosin (H&E) staining. Immunohistochemistry was performed to detect IL-9-positive (IL-9+) cells in lung tissues. Expression levels of IL-9, sIL-9R, STAT3, and p-STAT3 in peripheral blood of mice were determined by quantitative Real time-polymerase chain reaction (qRT-PCR), enzyme-linked immunosorbent assay (ELISA), and Western blot, respectively. In addition, the expression levels of superoxide dismutase (SOD), malondialdehyde (MDA), and reactive oxygen species (ROS) were detected. RESULTS: H&E staining results showed that the airway wall structure of COPD mice in the PBS group was irregular. Ciliated columnar epithelium exhibited marked degeneration, necrosis and shedding. Besides, numerous inflammatory cell infiltration, narrowing and rupture of the alveolar septa, and larger cysts fused by adjacent alveoli were observed. H&E staining also indicated that the structure of alveolar epithelium was severely impaired in COPD mice. However, the pathological changes in lung tissues of mice in the IL-9 antibody group were much milder than those of the PBS group. Immunohistochemistry results showed a significant deposition of IL-9+ cells in the lung tissues of the PBS group. Meanwhile, the mRNA and protein levels of IL-9, sIL-9R, and p-STAT3 in the PBS group were also remarkably higher than those of the IL-9 antibody group. In addition, SOD content in the PBS group was significantly decreased, whereas the levels of MDA and ROS were significantly increased than those of the IL-9 antibody group. CONCLUSIONS: IL-9 activated STAT3 and aggravated lung injury in COPD mice by increasing inflammatory and oxidative stress.

The therapeutic efficacy of pediatric ALL patients with MLL gene rearrangement treated with CCLG-ALL2008 protocol.

OBJECTIVE: In this study, we retrospectively evaluated the therapeutic efficacy of China Children Leukemia Group-ALL2008 (CCLG-ALL 2008) protocol in pediatric patients with mixed-lineage leukaemia (MLL) gene rearrangement of acute lymphoblastic leukemia (ALL) to identify the prognostic factors. PATIENTS AND METHODS: Six hundred and thirty-four patients with ALL were enrolled in this study between June 2008 and Dec 2014. High-risk group (HR) consisted of 217 cases, of which 28 cases were MLL related positive (first group), 22 cases were BCR/ABL positive (second group), and 167 cases were negative with MLL related or BCR/ABL (third group). The therapeutic efficacy was evaluated at the time points of day 8 (TP1), day 15 (TP2), day 33 (TP3) and 12th week (TP4) with the protocol, respectively. Overall-survival (OS) and relapse-free-survival (RFS) and treatment-related mortality (TRD) were analyzed as well. RESULTS: The first group accounted for 4.4% of all patients. Compared with the second and third group, the first group had more cases younger than 2 years, with initial leukocytes ≥50×109/L, and poor response on TP2. Moreover, patients older than 2 years old had a good 5 years OS (84% ± 9% vs. 37% ± 20%, p<0.05) and RFS (84% ± 9% vs. 29% ± 17%, p<0.05). There were no significant differences in the recurrence rate, TRD, 5 years OS and RFS among three groups. For the first group, compared with good response to prednisone, patients with poor response to prednisone had a poor 5 years RFS (41% ± 17% vs. 81% ± 10%, p<0.05). Multivariate Cox regression analysis identified that RFS and OS were influenced by such factors as age, MLL fusion partners, and prednisone response (p<0.05). CONCLUSIONS: Such factors as younger age than 2 years old, MLL/AF4 fusion gene, poor response to prednisone, or no complete remission (CR) on TP3 were poor prognostic parameters in predicting the outcome in childhood ALL with MLL gene rearrangement treated with CCLG-ALL 2008 protocol.

Suppressed CCL2 expression inhibits the proliferation of leukemia cells via the cell cycle protein Cyclin D1: preliminary in vitro data.

OBJECTIVE: Chemokine (C-C motif) ligand 2 (CCL2) is a member of the CC subfamily, which displays chemotactic activity for monocytes and basophils. This molecule plays a very important role in many solid tumors and shows changes in the bone marrow microenvironment. However, its role in acute myeloid leukaemia (AML) is still unclear. MATERIALS AND METHODS: In this study, we established a HL-60 cell line with CCL2 knockdown to explore its effect on leukemogenesis. Lentivirus with CCL2-knockdown was successfully constructed after screening effective CCL2 short hairpin RNA (shRNA) sequences and was transfected into HL-60 cells, which was further validated at the mRNA and protein levels by real-time polymerase chain reaction (PCR) and Western blotting, respectively. RESULTS: Low expression of CCL2 significantly decreased HL-60 cell growth by increasing the cell arrest at G1 phase by 12% more than controls. We applied RNA sequencing technology to discriminate the gene expression profiles between the cells with CCL2 knockdown and the controls, and Cyclin D1 was selected for further experiments as its expression level was significantly downregulated, which was validated at the mRNA and protein levels. Cyclin D1 knockdown experiments showed that the cell proliferation rate was evidently decelerated, and cell cycle analysis also indicated a similar pattern for CCL2. CONCLUSIONS: Our study revealed that Cyclin D1 is an effector that mediates CCL2's function in cell proliferation by blocking cells at G1 phase.

Association between proton pump inhibitors and the risk of hepatocellular carcinoma.

BACKGROUND: Proton pump inhibitor (PPI) use has been reported to be associated with liver damage and might possibly be carcinogenic. AIMS: We examined whether long-term PPI use increases the risk of hepatocellular carcinoma (HCC) in patients without viral hepatitis B or C. METHODS: We conducted a nested case-control study in a cohort of patients without viral hepatitis in Taiwan from 2000 to 2013. In total, 29 473 HCC cases and 294 508 matched controls were included. Moreover, we identified prescriptions for PPI and durations between the PPI index date and cancer diagnosis date (or the corresponding date in controls). RESULTS: The adjusted odds ratio (AOR) for HCC associated with PPI use was 2.86 (95% confidence interval [CI], 2.69-3.04). Considering the use of PPIs determined according to cumulative defined daily dose (cDDD) subgroups, a dose-response effect was observed in patients exposed to 29-180, 181-240, 241-300, and 300+ cDDDs of PPIs. The AORs were 2.74 (95% CI, 2.57-2.93), 2.98 (95% CI, 2.50-3.56), 3.23 (95% CI, 2.59-4.02), and 3.43 (95% CI, 2.94-4.00) in the 29-180, 181-240, 241-300, and 300+ cDDD groups, respectively, compared with the 0-28 cDDD group. A sensitivity analysis revealed a consistent association between PPI use and the risk of HCC in subpopulations stratified by risk factors associated with HCC. CONCLUSIONS: This observational study demonstrated that PPIs might increase the risk of HCC.

Dextrose monohydrate as a non-animal sourced alternative diluent in high shear wet granulation tablet formulations.

The feasibility of dextrose monohydrate as a non-animal sourced diluent in high shear wet granulation (HSWG) tablet formulations was determined. Impacts of granulation solution amount and addition time, wet massing time, impeller speed, powder and solution binder, and dry milling speed and screen opening size on granule size, friability and density, and tablet solid fraction (SF) and tensile strength (TS) were evaluated. The stability of theophylline tablets TS, disintegration time (DT) and in vitro dissolution were also studied. Following post-granulation drying at 60 °C, dextrose monohydrate lost 9% water and converted into the anhydrate form. Higher granulation solution amounts and faster addition, faster impeller speeds, and solution binder produced larger, denser and stronger (less friable) granules. All granules were compressed into tablets with acceptable TS. Contrary to what is normally observed, denser and larger granules (at ≥21% water level) produced tablets with a higher TS. The TS of the weakest tablets increased the most after storage at both 25 °C/60% RH and 40 °C/75% RH. Tablet DT was higher for stronger granules and after storage. Tablet dissolution profiles for 21% or less water were comparable and did not change on stability. However, the dissolution profile for tablets prepared with 24% water was slower initially and continued to decrease on stability. The results indicate a granulation water amount of not more than 21% is required to achieve acceptable tablet properties. This study clearly demonstrated the utility of dextrose monohydrate as a non-animal sourced diluent in a HSWG tablet formulation.

Carcinoembryonic antigen-related cell adhesion molecule 6 (CEACAM6) promotes EGF receptor signaling of oral squamous cell carcinoma metastasis via the complex N-glycosylation.

Aberrant protein glycosylation could be a distinct surface-marker of cancer cells that influences cancer progression and metastasis because glycosylation can regulate membrane protein folding which alters receptor activation and changes epitope exposure for antibody (Ab) recognition. Carcinoembryonic antigen-related cell adhesion molecule 6 (CEACAM6), a glycophosphoinositol-anchored protein, is a heavily glycosylated tumor antigen. However, the clinical significance and biological effect of CEACAM6 glycosylation has not been addressed in cancers. We recently developed an anti-CEACAM6 Ab (TMU) from an immune llama library which can be engineered to a single-domain (sd)Ab or a heavy-chain (HC)Ab. The TMU HCAb specifically recognized glycosylated CEACAM6 compared to the conventional antibodies. Using the TMU HCAb, we found that glycosylated CEACAM6 was a tumor marker associated with recurrence in early-stage OSCC (oral squamous cell carcinoma) patients. CEACAM6 promoted OSCC cell invasion, migration, cytoskeletal rearrangement, and metastasis via interaction with epidermal growth factor (EGF) receptor (EGFR) and enhancing EGFR activation, clustering and intracellular signaling cascades. These functions were modulated by N-acetylglucosaminyltransferase 5 (MGAT5) which mediated N-glycosylation at Asn256 (N256) of CEACAM6. Finally, the TMU sdAb and HCAb treatment inhibited the migration, invasion and EGF-induced signaling in CEACAM6-overexpressing cells. In conclusion, the complex N-glycosylation of CEACAM6 is critical for EGFR signaling of OSCC invasion and metastasis. Targeting glycosylated CEACAM6 with the TMU sdAb or TMU HCAb could be a feasible therapy for OSCC.

Feasibility of mini-tablets as a flexible drug delivery tool.

Mini-tablets have potential applications as a flexible drug delivery tool in addition to their generally perceived use as multi-particulates. That is, mini-tablets could provide flexibility in dose finding studies and/or allow for combination therapies in the clinic. Moreover, mini-tablets with well controlled quality attributes could be a prudent choice for administering solid dosage forms as a single unit or composite of multiple mini-tablets in patient populations with swallowing difficulties (e.g., pediatric and geriatric populations). This work demonstrated drug substance particle size and concentration ranges that achieve acceptable mini-tablet quality attributes for use as a single or composite dosage unit. Immediate release and orally disintegrating mini-tablet formulations with 30µm to 350µm (particle size d90) acetaminophen and Compap™ L (90% acetaminophen) at concentrations equivalent to 6.7% and 26.7% acetaminophen were evaluated. Mini-tablets achieved acceptable weight variability, tensile strength, friability, and disintegration time at a reasonable solid fraction for each formulation. The content uniformity was acceptable for mini-tablets of 6.7% formulations with ≤170µm drug substance, mini-tablets of all 26.7% formulations, and composite dosage units containing five or more mini-tablets of any formulation. Results supported the manufacturing feasibility of quality mini-tablets, and their applicability as a flexible drug delivery tool.

Cigarette smoking increases bronchial mucosal IL-17A expression in asthmatics, which acts in concert with environmental aeroallergens to engender neutrophilic inflammation.

BACKGROUND: Mild asthmatics who smoke cigarettes may develop unstable disease and neutrophilic infiltration of the airways, features more usually associated with severe asthmatic disease. The mechanisms giving rise to this response remain unclear. OBJECTIVE: To address the hypothesis that smoking increases bronchial mucosal production of IL-17A which acts on bronchial epithelial cells directly and in concert with other environmental stimuli to induce the production of IL-6 and neutrophil chemotaxins. METHODS: IL-17A, IL-8, IL-6, neutrophils and eosinophils were detected and quantified by immunohistochemistry in endobronchial biopsy sections from smoking and non-smoking asthmatics. Human tracheal epithelial cells (HTEpC) were cultured with IL-17A in the presence/absence of cigarette smoke extract (CSE) and aeroallergens lacking intrinsic protease activity, and IL-6 and IL-8 production measured in vitro. RESULTS: Expression of IL-17A, IL-6 and IL-8 and neutrophil numbers was significantly elevated in the bronchial mucosa of the asthmatic smokers compared to the non-smokers. Expression of IL-17A correlated with that of IL-8 and neutrophil numbers. In the smoking asthmatics, eosinophil numbers also correlated with expression of IL-8 and IL-17A. Exposure of HTEpC cells to both CSE and IL-17A increased expression of IL-6 and IL-8 in a concentration-dependent and synergistic manner. Co-stimulation with CSE, IL-17A and aeroallergens further increased IL-6 and IL-8 production synergistically. CONCLUSIONS: The data support the hypothesis that asthmatic smokers develop neutrophilic inflammation of the airways propagated at least partly by smoke-induced production of IL-17A which together with smoke and other environmental stimuli acts on airways epithelial cells to induce neutrophil chemotaxins.

Association between Anxiety and Pain in Dental Treatment: A Systematic Review and Meta-analysis.

Accumulating evidence has revealed that dental anxiety (DA), as a dispositional factor toward the dental situation, is associated with the state anxiety (SA) and pain related to dental procedures. However, conclusions from individual studies may be limited by the treatment procedures that patients received, the tools used to assess DA, or the treatment stages when anxiety or pain was assessed. It is unclear whether DA, at the study level, accounts for the variance in pretreatment SA. The impact of DA and SA on pain at different treatment stages has not been systematically investigated. To address these questions, we present novel meta-analytical evidence from 35 articles (encompassing 47 clinical groups) that investigated DA in a clinical group. Subgroup analyses revealed that the studies of surgical and nonsurgical procedures did not significantly differ in either DA or pretreatment SA. Furthermore, metaregressions revealed DA as a significant predictor that explained the variance in SA assessed before and during treatment but not after treatment. The findings suggest that patient DA has a significant impact on patient SA. Metaregressions revealed DA as a significant predictor that explained the variance in expected pain, pain during treatment and posttreatment pain. In contrast, pretreatment SA was a significant predictor that explained the variance in expected pain. The findings reveal that DA has a consistent impact on pain through the entire period of dental treatment. Altogether, the findings highlight the role of DA as an overall indicator for anxiety and pain, across different types of dental procedures or treatment stages. We conclude that anxiety should be assessed as a critical step not only in anxiety management for high-DA patients, but also in pain control for all dental patients.

Hypertension impairs hippocampus-related adult neurogenesis, CA1 neuron dendritic arborization and long-term memory.

Hypertension is associated with neurodegenerative diseases and cognitive impairment. Several studies using spontaneous hypertensive rats to study the effect of hypertension on memory performance and adult hippocampal neurogenesis have reached inconsistent conclusions. The contradictory findings may be related to the genetic variability of spontaneous hypertensive rats due to the conventional breeding practices. The objective of this study is to examine the effect of hypertension on hippocampal structure and function in isogenic mice. Hypertension was induced by the '2 kidneys, 1 clip' method (2K1C) which constricted one of the two renal arteries. The blood pressures of 2K1C mice were higher than the sham group on post-operation day 7 and remained high up to day 28. Mice with 2K1C-induced hypertension had impaired long-term, but not short-term, memory. Dendritic complexity of CA1 neurons and hippocampal neurogenesis were reduced by 2K1C-induced hypertension on post-operation day 28. Furthermore, 2K1C decreased the levels of hippocampal brain-derived neurotrophic factor, while blood vessel density and activation status of astrocytes and microglia were not affected. In conclusion, hypertension impairs hippocampus-associated long-term memory, dendritic arborization and neurogenesis, which may be caused by down-regulation of brain-derived neurotrophic factor signaling pathways.

Morphological characterization and molecular mapping of an irradiation-induced Speckled mutant in the silkworm, Bombyx mori.

Speckled (Spc), an X-ray-induced lethal mutant of Bombyx mori, exhibits a mosaic dark-brown-spotted larval epidermis in both sexes and egg-laying problems only in females. Here, we report the morphological characterization and molecular mapping of the Spc mutant. Morphological investigations revealed that the epidermal ultrastructure of the small, dark-brown spots was more dense than that of the white regions in both Spc/+ mutants and wild type, and that the lethality of the Spc/Spc mutants occurred during early embryogenesis. Furthermore, the ovarioles and ovipositor were disconnected in approximately 85.5% of Spc/+ females, a further 2.5% had a connection between the ovarioles and ovipositor that was too narrow to lay eggs. The remaining females showed a normal connection similar to that of the wild type. We successfully narrowed down the location of the Spc mutation to a region on chromosome 4 that was ∼1041 kb long. Gene-prediction analysis identified 25 candidate genes in this region. Chromosome structure analysis indicated that a ∼305 kb deletion was included in the mapping region. Temporal and spatial reverse transcription PCR (RT-PCR) analysis showed that several genes in the mapped region are associated with the Spc mutant. Although the genes responsible for the Spc mutation were not definitively identified, our results further the current understanding of the complex mechanism underlying the multiple morphological defects in Spc mutants.

Chirp- and random-based coded ultrasonic excitation for localized blood-brain barrier opening.

Chirp- and random-based coded excitation methods have been proposed to reduce standing wave formation and improve focusing of transcranial ultrasound. However, no clear evidence has been shown to support the benefits of these ultrasonic excitation sequences in vivo. This study evaluates the chirp and periodic selection of random frequency (PSRF) coded-excitation methods for opening the blood-brain barrier (BBB) in mice. Three groups of mice (n = 15) were injected with polydisperse microbubbles and sonicated in the caudate putamen using the chirp/PSRF coded (bandwidth: 1.5­1.9 MHz, peak negative pressure: 0.52 MPa, duration: 30 s) or standard ultrasound (frequency: 1.5 MHz, pressure: 0.52 MPa, burst duration: 20 ms, duration: 5 min) sequences. T1-weighted contrast-enhanced MRI scans were performed to quantitatively analyze focused ultrasound induced BBB opening. The mean opening volumes evaluated from the MRI were mm3, mm3and mm3 for the chirp, random and regular sonications, respectively. The mean cavitation levels were V.s, V.s and V.s for the chirp, random and regular sonications, respectively. The chirp and PSRF coded pulsing sequences improved the BBB opening localization by inducing lower cavitation levels and smaller opening volumes compared to results of the regular sonication technique. Larger bandwidths were associated with more focused targeting but were limited by the frequency response of the transducer, the skull attenuation and the microbubbles optimal frequency range. The coded methods could therefore facilitate highly localized drug delivery as well as benefit other transcranial ultrasound techniques that use higher pressure levels and higher precision to induce the necessary bioeffects in a brain region while avoiding damage to the surrounding healthy tissue.

Mechanism for enhanced absorption of a solid dispersion formulation of LY2300559 using the artificial stomach duodenum model.

An artificial stomach duodenum (ASD) model has been used to demonstrate the performance difference between two formulations of LY2300559, a low-solubility acidic developmental drug. The two formulations investigated were a conventional high-shear wet granulation (HSWG) formulation and a solid dispersion formulation. A pharmacokinetic study in humans demonstrated the enhanced performance of the solid dispersion formulation relative to the HSWG formulation. The Cmax and AUC of the solid dispersion was 2.6 and 1.9 times greater, respectively, compared to the HSWG formulation. In the ASD, the solid dispersion formulation performance was characterized by three main phases: (1) rapid release in the stomach, creating a supersaturated concentration of drug, (2) precipitation in the stomach, and (3) rapid redissolution of the precipitate in the duodenum to concentration levels that are supersaturated relative to crystalline drug. A series of complementary experiments were employed to describe this performance behavior mechanistically. Imaging experiments with a pH indicating dye showed that local pH gradients from meglumine in the solid dispersion formulation were responsible for creating a high initial supersaturation concentration in the stomach. Upon dissipation of meglumine, the drug precipitated in the stomach as an amorphous solid. Because the precipitated drug is in an amorphous form, it can then rapidly redissolve as it transits to the more neutral environment of the duodenum. This unexpected sequence of physical state changes gives a mechanistic explanation for the enhanced in vivo performance of the solid dispersion formulation relative to the HSWG formulation.

A detailed analysis of the leaf rolling mutant sll2 reveals complex nature in regulation of bulliform cell development in rice (Oryza sativa L.).

Bulliform cells are large, thin-walled and highly vacuolated cells, and play an important role in controlling leaf rolling in response to drought and high temperature. However, the molecular mechanisms regulating bulliform cell development have not been well documented. Here, we report isolation and characterisation of a rice leaf-rolling mutant, named shallot-like 2 (sll2). The sll2 plants exhibit adaxially rolled leaves, starting from the sixth leaf stage, accompanied by increased photosynthesis and reduced plant height and tiller number. Histological analyses showed shrinkage of bulliform cells, resulting in inward-curved leaves. The mutant is recessive and revertible at a rate of 9%. The leaf rolling is caused by a T-DNA insertion. Cloning of the insertion using TAIL-PCR revealed that the T-DNA was inserted in the promoter region of LOC_Os07 g38664. Unexpectedly, the enhanced expression of LOC_Os07 g38664 by the 35S enhancer in the T-DNA is not responsible for the leaf rolling phenotype. Further, the enhancer also exerted a long-distance effect, including up-regulation of several bulliform cell-related genes. sll2 suppressed the outward leaf rolling of oul1 in the sll2oul1 double mutant. We conclude that leaf rolling in sll2 could be a result of the combined effect of multi-genes, implying a complex network in regulation of bulliform cell development.

Genetic polymorphisms in the prostaglandin pathway genes and risk of head and neck cancer.

OBJECTIVE: Previous studies examining the association between genetic variations in prostaglandin pathway and risk of head and neck cancer (HNC) have only included polymorphisms in the PTGS2 (COX2) gene. This study investigated the association between genetic polymorphisms of six prostaglandin pathway genes (PGDS, PTGDS, PTGES, PTGIS, PTGS1 and PTGS2), and risk of HNC. METHODS: Interviews regarding the consumption of alcohol, betel quid, and cigarette were conducted with 222 HNC cases and 214 controls. Genotyping was performed for 48 tag and functional single-nucleotide polymorphisms (SNPs). RESULTS: Two tag SNPs of PTGIS showed a significant association with HNC risk [rs522962: log-additive odds ratio (OR) = 1.42, 95% confidence interval (CI): 1.01-1.99 and dominant OR = 1.58, 95% CI: 1.02-2.47; rs6125671: log-additive OR = 1.49, 95% CI: 1.08-2.05 and dominant OR = 1.96, 95% CI: 1.16-3.32]. In addition, a region in PTGIS tagged by rs927068 and rs6019902 was significantly associated with risk of HNC (global P = 0.007). Finally, several SNPs interacted with betel quid and cigarette to influence the risk of HNC. CONCLUSIONS: Genetic variations in prostaglandin pathway genes are associated with risk of HNC and may modify the relationship between use of betel quid or cigarette and development of HNC.

MicroRNA-17-5p post-transcriptionally regulates p21 expression in irradiated betel quid chewing-related oral squamous cell carcinoma cells.

BACKGROUND AND PURPOSE: Betel nut chewing is associated with oral cavity cancer in Taiwan. OC3 is an oral carcinoma cell line that was established from cells collected from a long-term betel nut chewer who does not smoke. After we found that microRNA-17-5p (miR-17-5p) is induced in OC3 cells, we used this cell line to examine the biological role(s) of this microRNA in response to exposure to ionizing radiation. MATERIALS AND METHODS: A combined SYBR green-based real-time PCR and oligonucleotide ligation assay was used to examine the expression of the miR-17 polycistron in irradiated OC3 cells. The roles of miR-17-5p and p21 were evaluated with specific antisense oligonucleotides (ODN) that were designed and used to inhibit their expression. Expression of the p21 protein was evaluated by Western blotting. The clonogenic assay and annexin V staining were used to evaluate cell survival and apoptosis, respectively. Cells in which miR-17-5p was stably knocked down were used to create ectopic xenografts to evaluate in vivo the role of miR-17-5p. RESULTS: A radiation dose of 5 Gy significantly increased miR-17-5p expression in irradiated OC3 cells. Inhibition of miR-17-5p expression enhanced the radiosensitivity of the OC3 cells. We found that miR-17-5p downregulates radiation-induced p21 expression in OC3 cells and, by using a tumor xenograft model, it was found that p21 plays a critical role in increasing the radiosensitivity of OC3 cells in vitro and in vivo. CONCLUSION: miR-17-5p is induced in irradiated OC3 cells and it downregulates p21 protein expression, contributing to the radioresistance of OC3 cells.

The HPV E6 oncoprotein targets histone methyltransferases for modulating specific gene transcription.

Expression of viral proteins causes important epigenetic changes leading to abnormal cell growth. Whether viral proteins directly target histone methyltransferases (HMTs), a key family enzyme for epigenetic regulation, and modulate their enzymatic activities remains elusive. Here we show that the E6 proteins of both low-risk and high-risk human papillomavirus (HPV) interact with three coactivator HMTs, CARM1, PRMT1 and SET7, and downregulate their enzymatic activities in vitro and in HPV-transformed HeLa cells. Furthermore, these three HMTs are required for E6 to attenuate p53 transactivation function. Mechanistically, E6 hampers CARM1- and PRMT1-catalyzed histone methylation at p53-responsive promoters, and suppresses the binding of p53 to chromatinized DNA independently of E6-mediated p53 degradation. p53 pre-methylated at lysine-372 (p53K372 mono-methylation) by SET7 protects p53 from E6-induced degradation. Consistently, E6 downregulates p53K372 mono-methylation and thus reduces p53 protein stability. As a result of the E6-mediated inhibition of HMT activity, expression of p53 downstream genes is suppressed. Together, our results not only reveal a clever approach for the virus to interfere with p53 function, but also demonstrate the modulation of HMT activity as a novel mechanism of epigenetic regulation by a viral oncoprotein.