Changes in acetylcholinesterase activity and muscarinic receptor bindings in mu-opioid receptor knockout mice

Anatomical evidence indicates that cholinergic and opioidergic systems are co-localized and acting on the same neurons. However, the regulatory mechanisms between cholinergic and opioidergic system have not been well characterized. In the present study, we investigated whether there are compensatory changes of acetylcholinesterase activity and cholinergic receptors in mice lacking mu-opioid receptor gene. The acetylcholinesterase activity was determined by histochemistry assay. The cholinergic receptor binding was carried out by quantitative autoradiography using [3H]-quinuclidinyl benzilate (nonselective muscarinic receptors), N-[3H]-methylscopolamine (nonselective muscarinic receptors), [3H]-pirenzepine (M1 subtype muscarinic receptors) and [3H]-AF-DX384 (M2 subtype muscarinic receptors) in brain slices of wild-type and mu-opioid receptor knockout mice. The acetylcholinesterase activity of mu-opioid receptor knockout mice was higher than that of the wild-type in the striatal caudate putamen and nucleus accumbens, but not in the cortex and hippocampus areas. In addition, the bindings in N-[3H]-methylscopolamine and [3H]-AF-DX384 of mu-opioid receptor knockout mice were significantly lower when compared with that of the wild-type controls in the striatal caudate putamen and nucleus accumbens. However, there were no significant differences in bindings of [3H]-quinuclidinyl benzilate and [3H]-pirenzepine between mu-opioid receptor knockout and wild-type mice in the cortex, striatum and hippocampus. These data indicate that there are up-regulation of acetylcholinesterase activity and compensatory down-regulation of M2 muscarinic receptors in the striatal caudate putamen and nucleus accumbens of mu-opioid receptor knockout mice.     

Is retinal photography useful in the measurement of stroke risk?

BACKGROUND: The retinal microcirculation can be viewed non-invasively to give a unique perspective of the cerebral microcirculation in vivo. Studying pathological changes of retinal blood vessels (microaneurysms, retinal haemorrhages, and retinal arteriolar narrowing) may help to understand the causes of various cerebrovascular disorders. Retinal photography provides such an opportunity. RECENT DEVELOPMENTS: Several recent studies have shown that retinal microvascular changes are reliably documented by retinal photographs. These retinopathy changes seem to be fairly common in the general population, even in people without hypertension or diabetes. Retinopathy is related to incident clinical stroke and stroke mortality and to MRI-defined subclinical cerebral white-matter lesions and cerebral atrophy, independent of blood pressure, diabetes, and other cerebrovascular risk factors. WHERE NEXT?: Retinal microvascular abnormalities seem to be markers of concomitant cerebral microangiopathy, and retinal photography may be useful for the investigation of microvascular disorders of the brain in clinical and epidemiological settings. Future research should be aimed at the development of standardised photographic methods for the assessment of retinal microvascular changes, the replication of these findings in other populations and in people with other cerebrovascular disorders, and the examination of the increased accuracy of stoke-risk stratification given by retinal photography     

Does refractive error influence the association of blood pressure and retinal vessel diameters? The Blue Mountains Eye Study

PURPOSE: To determine if refractive errors influence the association of blood pressure and retinal vessel diameters. DESIGN: Population-based, cross-sectional study. METHODS: Retinal photographs from the right eyes of participants (n = 3,654, aged 49+ years) in the Blue Mountains Eye Study taken during baseline examinations (1992 to 1994) were digitized. The diameter of all retinal vessels located half to one disk diameter from the disk margin was measured using a computer-assisted imaging program. These measurements were combined to provide the average diameters of retinal arterioles and venules of that eye, and the ratio of their diameters, the arteriole-to-venule ratio (AVR). The association of blood pressure and retinal vessel diameters was analyzed before and after correction for refraction using the Bengtsson formula. RESULTS: Before correction, each 10-mm Hg increase in mean arterial blood pressure was associated with a 3.7-microm (95% confidence interval [CI], 3.2-4.3) decrease in arteriolar diameter and a 0.9-microm (95% CI, 0.3-0.9) decrease in venular diameter. After correction for refraction, each 1-mm Hg increase in mean arterial blood pressure was associated with a 3.7-microm (95% CI, 3.2-4.2) decrease in arteriolar diameter and a 0.8-microm (95% CI, 0.3-0.9) decrease in venular diameter. Refraction was not associated with the AVR and had no effect on the association of blood pressure and AVR. CONCLUSION: Refraction had no appreciable effect on the association of blood pressure and retinal vessel diameters or on the AVR. Correction for refraction is important for quantifying absolute retinal vessel caliber, but may not be particularly important in epidemiologic studies investigating the association of generalized retinal arteriolar narrowing and hypertension.     

Endometrial effects of lower doses of conjugated equine estrogens and medroxyprogesterone acetate: two-year substudy results

OBJECTIVE: To determine the endometrial safety of 2 years of treatment with lower doses of continuous combined conjugated equine estrogens (CEE) and medroxyprogesterone acetate (MPA). DESIGN: Randomized, double-blind, placebo-controlled, multicenter metabolic and osteoporosis substudy of the Women's Health, Osteoporosis, Progestin, Estrogen (Women's HOPE) study. SETTING: Nineteen study centers across the United States. PATIENT(S): Healthy, postmenopausal women (n = 822) with an intact uterus were recruited. INTERVENTION(S): Patients received CEE 0.625, CEE 0.625/MPA 2.5, CEE 0.45, CEE 0.45/MPA 2.5, CEE 0.45/MPA 1.5, CEE 0.3, CEE 0.3/MPA 1.5 (all doses mg/day), or placebo for 2 years. Endometrial biopsies were evaluated at baseline and years 0.5, 1, 1.5, and 2 using a centralized protocol. MAIN OUTCOME MEASURE(S): Efficacy of lower doses of CEE/MPA in reducing the incidence of endometrial hyperplasia rates associated with unopposed estrogen (E). RESULT(S): No cases of endometrial hyperplasia were seen in the four CEE/MPA groups. For the CEE-alone groups, a dose-related increase in incidence rates from 3.17% (CEE 0.3 mg) to 27.27% (CEE 0.625 mg) was seen at 2 years. The number of cases increased from year 1 to year 2. For the CEE-alone groups, the incidence rates and types of hyperplasia diagnosed varied among the pathologists. CONCLUSION(S): Two years of treatment with lower doses of CEE/MPA provided endometrial protection comparable to that seen with commonly prescribed doses. These regimens should be considered for postmenopausal women who are candidates for hormone therapy.     

Repositioning of cells by mechanotaxis on surfaces with micropatterned Young's modulus

Adherent cells are strongly influenced by the mechanical aspects of biomaterials, but little is known about the cellular effects of spatial variations in these properties. This work describes a novel method to produce polymeric cell culture surfaces containing micrometer-scale regions of variable stiffness. Substrates made of acrylamide or poly(dimethylsiloxane) were patterned with 100- or 10-microm resolution, respectively. Cells were cultured on fibronectin-coated acrylamide having Young's moduli of 34 kPa and 1.8 kPa, or fibronectin-coated PDMS having moduli of 2.5 MPa and 12 kPa. Over several days, NIH/3T3 cells and bovine pulmonary arterial endothelial cells accumulated preferentially on stiffer regions of substrates. The migration, not proliferation, of cells in response to mechanical patterning (mechanotaxis) was responsible for the accumulation of cells on stiffer regions. Differential remodeling of extracellular matrix protein on stiff versus compliant regions was observed by immunofluorescence staining, and may have been responsible for the observed mechanotaxis. These results suggest that mechanically patterned substrates might provide a general means to study mechanotaxis, and a new approach to patterning cells.     

A composite graft material containing bone particles and collagen in osteoinduction in mouse.

Demineralized allogenic bone matrices (DABM) and demineralized freeze-dried bone allograft (DFDBA) have been successfully used as bone-graft materials in the treatment of acquired and congenital cranio-maxillofacial defects and in some orthopedic surgery. However, these bone-graft "powders" have many shortcomings. For example, placement of particulate graft material in a hemorrhaging site can result in inadequacies or inaccurate attachment as well as loss of the graft materials. To minimize the inadequacies of powderlike graft materials, xenogenic collagen isolated from human tendon, skin, or bone was added to the bone-graft particles to form a composite spongelike implant. This material is commercially available and consists of 60% collagen and 40% DFDBA (DynaGraft, GenSci Co., Irvine, CA). The goal of this study was to evaluate the characteristics of composite graft implants in the mineralization process in an animal model in comparison with DFDBA powder and pure collagen. Seventy-two Swiss Webster mice were divided into three groups: an experimental group implanted with DynaGraft, two comparison groups implanted with either DFDBA or collagen only. All the graft materials were surgically implanted and inserted into the left thigh muscle. Mice were humanely killed at 1, 2, 3, 4, 6, 8, and 12 weeks. Then the muscle tissues in the vicinity of the implants were excised and processed for histology. Paraffin sections were stained with hematoxylin and eosin (H&E), the Von Kossa method, and Masson's trichrome. Some selected specimens were processed for transmission electron microscopic observation. After 1 week of implantation, the DynaGraft group showed calcium deposition on the collagen material and on the periphery of the DFDBA particles. Increased calcification and bone-forming cells were observed at 4-6 weeks. After 8 weeks, the implant formed a calcified nodule and only heavily mineralized connective tissue was observed at the implanted site. The group implanted with DFDBA powder showed calcification around the particulates. The collagen-sponge control group revealed no calcification or bone formation during the period of implantation. The light microscopic findings were confirmed by electron microscopy. Quantitative radiographic density DynaGraft and DFDBA graft followed sequentially over a period 120 days. It was concluded that a higher rate of calcification and bone formation was produced in the composite graft implant compared to the DFDBA implant. The composite graft material (DynaGraft), which contains both collagen and DFDBA, proved to be more effective for bone formation than particle components alone.     

Control of aedes vectors of dengue in three provinces of Vietnam by use of Mesocyclops (Copepoda) and community-based methods validated by entomologic,clinical, and serological surveillance

We describe remarkable success in controlling dengue vectors, Aedes aegypti (L.) and Aedes albopictus (Skuse), in 6 communes with 11,675 households and 49,647 people in the northern provinces of Haiphong, Hung Yen, and Nam Dinh in Vietnam. The communes were selected for high-frequency use of large outdoor concrete tanks and wells. These were found to be the source of 49.6-98.4% of Ae. aegypti larvae, which were amenable to treatment with local Mesocyclops, mainly M. woutersi Van der Velde, M. aspericornis (Daday) and M. thermocyclopoides Harada. Knowledge, attitude, and practice surveys were performed to determine whether the communities viewed dengue and dengue hemorrhagic fever as a serious health threat; to determine their knowledge of the etiology, attitudes, and practices regarding control methods including Mesocyclops; and to determine their receptivity to various information methods. On the basis of the knowledge, attitude, and practice data, the community-based dengue control program comprised a system of local leaders, health volunteer teachers, and schoolchildren, supported by health professionals. Recycling of discards for economic gain was enhanced, where appropriate, and this, plus 37 clean-up campaigns, removed small containers unsuitable for Mesocyclops treatment. A previously successful eradication at Phan Boi village (Hung Yen province) was extended to 7 other villages forming Di Su commune (1,750 households) in the current study. Complete control was also achieved in Nghia Hiep (Hung Yen province) and in Xuan Phong (Nam Dinh province); control efficacy was > or = 99.7% in the other 3 communes (Lac Vien in Haiphong, Nghia Dong, and Xuan Kien in Nam Dinh). Although tanks and wells were the key container types of Ae. aegypti productivity, discarded materials were the source of 51% of the standing crop of Ae. albopictus. Aedes albopictus larvae were eliminated from the 3 Nam Dinh communes, and 86-98% control was achieved in the other 3 communes. Variable dengue attack rates made the clinical and serological comparison of control and untreated communes problematic, but these data indicate that clinical surveillance by itself is inadequate to monitor dengue transmission.     

Retinal vascular abnormalities in persons with type 1 diabetes: the Wisconsin Epidemiologic Study of Diabetic Retinopathy: XVIII

PURPOSE: To describe the distribution of retinal vascular characteristics and their correlates in people with type 1 diabetes. DESIGN: Population-based study with baseline cross-sectional findings. PARTICIPANTS: Nine hundred ninety-six persons who were diagnosed with diabetes before 30 years of age and who were taking insulin in an 11-county area in south-central Wisconsin participated in the baseline examination from 1980 to 1982. In addition, 225 persons without diabetes participated. METHODS: Retinal photographs of 7 standard fields were taken; lightbox grading was performed to determine arteriovenous (A/V) nicking and focal retinal arteriolar narrowing. Computer-assisted grading was performed from a digitized image of field 1 to determine central retinal arteriolar equivalent (CRAE), central retinal venular equivalent (CRVE), and the arteriole-to-venule ratio (AVR). MAIN OUTCOME MEASURES: Frequency and distribution of A/V nicking, CRAE, CRVE, AVR, and focal retinal arteriolar narrowing. RESULTS: In multivariate analyses, retinopathy severity, mean arterial blood pressure, presence of gross proteinuria, glycosylated hemoglobin, and history of cardiovascular disease were associated with CRAE. Retinopathy severity, age, mean arterial blood pressure, duration of diabetes, glycosylated hemoglobin level, and body mass index were associated with CRVE. With the exception of glycosylated hemoglobin, similar factors were associated with AVR. Age (odds ratio [OR] per 10 years, 2.43 and 2.02) and retinopathy severity (OR per level, 1.14 and 1.21) were associated with focal retinal arteriolar narrowing and A/V nicking, respectively. In persons >/=18 years of age, smoking was associated with CRAE, CRVE, and A/V nicking (OR, 2.67), but not with AVR or focal arteriolar narrowing. CONCLUSIONS: This study documented the frequency and distribution of retinal vascular characteristics and their relationships to various factors in persons with type 1 diabetes.     

Inhibitory effect of kaurane type diterpenoids from<Emphasis Type="Italic">Acanthopanax koreanum</Emphasis> on TNF-α secretion from trypsin-stimulated HMC-1 cells

Five known kaurane type diterpenoids, 16alphaH,17-isovaleryloxy-ent-kauran-19-oic acid (1), 16alpha-hydroxy-17-isovaleryloxy-ent-kauran-19-oic acid (2), paniculoside-IV (3), 16alpha-hydroxy-ent-kauran-19-oic acid (4), and ent-kaur-16-en-19-oic acid (5) were isolated from the root of Acanthopanax koreanum by repeated column chromatography and reversed phase preparative HPLC. The structures of these compounds were established from physicochemical and spectral data. Among the isolated compounds 16alphaH,17-isovaleryloxy-ent-kauran-19-oic acid (1) showed potent inhibitory activity (IC50 value, 16.2 uM) on TNF-alpha secretion from HMC-1, a trypsin-stimulated human leukemic mast cell line.     

Overexpression of mutated MRP4 in cisplatin resistant small cell lung cancer cell line: collateral sensitivity to azidothymidine

Cisplatin (CDDP) resistance is one of the major impediments in cancer chemotherapy. In an attempt to define this complex mechanism(s) of resistance, we have identified 7 cDNA fragments which are overexpressed in CDDP resistant small cell lung cancer cell line (SR-2) using PCR selected cDNA subtraction. One of these fragments was identical with nucleotide 3657-4042 of MRP4. The other fragments share sequence homology with elongation factor alpha, human placenta villi cDNA, heat shock protein (Hsp70), ribosomal RNA, BNP1 brain specific Na-dependent inorganic phosphate cotransporter and telomeric catalytic subunit. Examination of other MRP members (MRP1, 2, 3, 5, 6) did not show discernable differences in their expression between the parental (SCLC1) and the CDDP-resistant variant (SR-2). Full length MRP4 cDNA was obtained from SCLC1 and SR-2. Both cell lines carry a point mutation at nucleotide 3532 while SR-2 carries two additional mutations at 3228 and 3246. Since MRP4 is known to transport azidiothymidine (AZT) and overexpression of MRP4 confers AZT resistance, we have studied growth inhibitory effects of AZT and [3H]-AZT accumulation. Interestingly, SR-2 is more sensitive to AZT while accumulating lesser amounts of [3H]-AZT. The thymidine kinase activity is similar in both cell lines. Thus, the increased sensitivity to AZT in SR-2 could not be solely due to mutation of MRP4. These findings are most likely due to the inhibitory effects of telomere catalytic subunit by AZT. Thus, certain biochemical changes induced by CDDP can be explored for future treatment to overcome this form of resistance.