[Clinical application of photobiomodulation in trauma repair and medical aesthetics].

In recent years, with the deepening of researches on the molecular biological mechanisms of photobiomodulation (PBM), PBM has gradually been applied in clinical practice, providing effective treatment methods and approaches for various diseases. Compared with traditional photothermal therapy, PBM has the characteristics of good therapeutic effect, almost no adverse reaction, and simple operation, and its clinical efficacy is becoming increasingly significant. This article provides a detailed explanation on the mechanism of PBM, its application characteristics and development trends in trauma repair and medical aesthetics, in order to provide a theoretical basis for the extensively clinical application of this therapy.

[Prospective randomized controlled trial on 2 940 nm fractional erbium laser combined with fractional micro-plasma radiofrequency in the treatment of facial atrophic acne scars].

Objective: To compare the efficacy and safety of 2 940 nm fractional erbium laser combined with fractional micro-plasma radiofrequency (FMR) therapy and 2 940 nm fractional erbium laser in the treatment of atrophic acne scars. Methods: A prospective randomized controlled research was conducted. A total of 100 atrophic acne scar patients (38 males and 62 females, aged 18-37 years) who were treated in the Scar Laser Clinic of the Department of Plastic and Reconstructive Surgery of Shanghai Ninth People's Hospital of Shanghai Jiao Tong University School of Medicine from March 2018 to March 2021 and conformed to the inclusion criteria were recruited. The patients were randomly divided into erbium laser+FMR group and erbium laser alone group, with 50 cases in each group. The facial acne scars of patients in erbium laser alone group were treated with 2 940 nm fractional erbium laser, while the facial acne scars of patients in erbium laser+FMR group were treated with erbium laser as above, besides, the scars of U and M types were treated with FMR, once every 3 months for a total of 3 times. Before the first treatment and 3 months after each treatment, the Echelle D'Assessment Clinique des Cicatrices D'Acne (ECCA) was used to score the scar. The occurrence of adverse reaction during the treatment process was observed and recorded, and the incidence was calculated. Three months after the last treatment, the 5-level classification method was used to evaluate the satisfaction of patients with the treatment effect, and the satisfaction rate was calculated. Data were statistically analyzed with independent sample t test and chi-square test. Results: A total of 89 patients completed the study, including 46 patients in erbium laser+FMR group (19 males and 27 females, aged (26±5) years) and 43 patients in erbium laser alone group (15 males and 28 females, aged (27±6) years). The ECCA scores before the first treatment and 3 months after the first treatment of patients were similar between the two groups (P>0.05). The ECCA scores of patients in erbium laser+FMR group at 3 months after the second and third treatment were 72±23 and 61±18, respectively, which were significantly lower than 85±26 and 76±25 in erbium laser alone group (with t values of -2.45 and -3.26, respectively, P<0.05). During the treatment process, the incidence of adverse reaction of patients in erbium laser+FMR group and erbium laser alone group were 23.91% (11/46) and 16.28% (7/43), respectively, and there was no statistically significant difference between the two groups (P>0.05). The satisfaction rate of patients in erbium laser+FMR group was 78.26% (36/46) at 3 months after the last treatment, which was significantly higher than 53.49% (23/43) in erbium laser alone group (χ2=6.10, P<0.05). Conclusions: The 2 940 nm fractional erbium laser combined with FMR is superior to 2 940 nm fractional erbium laser alone in the treatment of facial atrophic acne scars, achieving significantly higher efficacy without significantly increasing the incidence of adverse reaction, and patients are more satisfied with the efficacy. It can be used as a recommended therapy in clinical practice.

[The association of Vitamin D levels with lipid metabolism during pregnancy].

Vitamin D deficiency is common in pregnant women, and it is associated with multiple gestational complications, fetal intrauterine dysplasia, and maternal and infant health in postpartum period. Vitamin D involved in lipid metabolism may affect maternal-fetal health during pregnancy through this biological pathway. This review provides an overview of three main aspects, including, the effect of vitamin D on the mechanism of lipid metabolism, the relationship between vitamin D and lipid metabolites in pregnant women and women with pregnancy complications. An association between vitamin D status and lipid metabolism at different stages of pregnancy has been justified in most current studies, and the underlying mechanism has also been partly elucidated. Given some gestational complications, such as gestational diabetes and pre-eclampsia, pregnant women with these conditions are often associated with abnormal lipid metabolism and vitamin D deficiency. There is no doubt that the studies on the vitamin D supplementation and lipid metabolism improvement could have crucial clinical significance for this population. More randomized controlled trials with multi-center, large sample and multi-dose interventions are needed in the future to provide scientific evidence for clinical practice.

[Clinical effects of a combination treatment with narrow-spectrum intense pulsed light and fractional carbon dioxide laser on hypertrophic scar pruritus].

Objective: To observe the effects of a combination treatment with narrow-spectrum intense pulsed light and fractional carbon dioxide laser on hypertrophic scar pruritus in clinic. Methods: A prospective randomized controlled study was conducted. A total of 90 patients with hypertrophic scars conforming to the inclusion criteria who were hospitalized in our ward from March to December 2017 were divided into combination treatment group and control group according to the random number table, with 45 patients in each group. For scar pruritus, patients in control group were treated twice by narrow-spectrum intense pulsed light with a one-month interval, while patients in combination treatment group were firstly treated by narrow-spectrum intense pulsed light combined with fractional carbon dioxide laser once, and with narrow-spectrum intense pulsed light once one month later. Before and 3 months after treatment, scar pruritus was evaluated by the Visual Analogue Scale and the Four-item Itch Questionnaire, and the improvement of scar was assessed by photography. Three months after treatment, the treatment satisfaction of scar pruritus was self-rated by patients. Meanwhile, adverse effects were recorded during the procedures and follow-up periods. Data were processed with Chi-square test, paired t test, Wilcoxon rank-sum test, and Fisher's exact probability test. Results: Before treatment, there was no statistically significant difference in scar pruritus degree evaluated by the Visual Analogue Scale and score distribution of scar pruritus location, extent, frequency, and sleep effect of the Four-item Itch Questionnaire between patients of two groups (Z=-1.08, -0.91, -0.03, -0.69, -1.49, P>0.05). Three months after treatment, there was reducing degree of scar pruritus of patients evaluated by the Visual Analogue Scale in control group or combination treatment group compared with before treatment within the same group (Z=-1.98, -4.65, P<0.05 or P<0.01), while the latter was more obvious than the former (Z=-2.14, P<0.05). There were reducing scores of scar pruritus extent and frequency of patients in control group, along with location, extent, frequency, and sleep effect of patients in combination treatment group compared with those before treatment within the same group (Z=-2.33, -2.34, -3.53, -4.96, -3.32, -4.84, P<0.05 or P<0.01). However, scores of scar pruritus location and sleep effect of patients in control group were similar to those before treatment within the same group (Z=-0.58, -1.34, P>0.05). The scores of scar pruritus location, extent, frequency, and sleep effect of patients were obviously lower in combination treatment group compared with control group (Z=-2.09, -2.69, -1.99, -2.23, P<0.05 or P<0.01). It was much better of scar improvement of patients in combination treatment group compared with control group (Z=-4.00, P<0.01). The percentages of treatment satisfaction of scar pruritus of patients with 0, 1%-25%, 26%-50%, 51%-75%, 76%-100% were 0, 2.2% (1/45), 17.8% (8/45), 48.9% (22/45), and 31.1% (14/45) respectively in combination treatment group, which were obviously better than 0, 11.1% (5/45), 53.3% (24/45), 28.9% (13/45), and 6.7% (3/45) in control group (Z=-4.42, P<0.01). During the treatment and follow-up periods, the adverse effect ratio of patients in control group was 6.7% (3/45), similar to 2.2% (1/45) of combination treatment group (P>0.05). Conclusions: The combination of narrow-spectrum intense pulsed light and fractional carbon dioxide laser can greatly reduce pruritus, improve effect of scar treatment, and bring higher patient satisfaction compared with narrow-spectrum intense pulsed light alone in treating hypertrophic scar pruritus. Clinical trial registration: Chinese Clinical Trial Registry, ChiCTR-ONH-17012350.

Impact of community-based integrated mass drug administration on schistosomiasis and soil-transmitted helminth prevalence in Togo.

BACKGROUND: Togo has conducted annual, integrated, community-based mass drug administration (MDA) for soil-transmitted helminths (STH) and schistosomiasis since 2010. Treatment frequency and target populations are determined by disease prevalence, as measured by baseline surveys in 2007 and 2009, and WHO guidelines. Reported programmatic treatment coverage has averaged over 94%. Togo conducted a cross-sectional survey in 2015 to assess the impact of four to five years of MDA on these diseases. METHODOLOGY/PRINCIPAL FINDINGS: In every sub-district in the country outside the capital, the same schools were visited as at baseline and a sample of fifteen children age 6 to 9 years old was drawn. Each child submitted urine and a stool sample. Urine samples were tested by dipstick for the presence of blood as a proxy measure of Schistosoma haematobium infection. Stool samples were analyzed by the Kato-Katz method for STH and Schistosoma mansoni. At baseline, 17,100 children were enrolled at 1,129 schools in 562 sub-districts; in 2015, 16,890 children were enrolled at the same schools. The overall prevalence of both STH and schistosomiasis declined significantly, from 31.5% to 11.6% for STH and from 23.5% to 5.0% for schistosomiasis (p<0.001 in both instances). Egg counts from both years were available only for hookworm and S. mansoni; intensity of infection decreased significantly for both infections from 2009 to 2015 (p<0.001 for both infections). In areas with high baseline prevalence, rebound of hookworm infection was noted in children who had not received albendazole in the past 6 months. CONCLUSIONS/SIGNIFICANCE: After four to five years of MDA in Togo, the prevalence and intensity of STH and schistosomiasis infection were significantly reduced compared to baseline. Data on STH indicate that stopping MDA in areas with high baseline prevalence may result in significant rebound of infection. Togo's findings may help refine treatment recommendations for these diseases.

Wogonin increases doxorubicin sensitivity by down-regulation of IGF-1R/AKT signaling pathway in human breast cancer.

Currently drug resistance has remained a major challenge in successful breast cancer therapy. Wogonin, one of the active components of scutellaria baicalensis, has shown anticarcinogenic, chemopreventive, and immunoregulatory functions. The present study aimed to explore whether wogonin exerted synergistic cytotoxicity with doxorubicin in breast cancer. Our data indicated that wogonin inhibited the proliferation of breast cancer cells in a dose- and time-dependent manner. Combined treatment with wogonin increased the doxorubicin sensitivity in breast cancer cells. Moreover, administration with wogonin alone or in combination with doxorubicin suppressed the expression of insulin like growth factor 1 receptor (IGF-1R) in Bcap-37 and MCF-7 cells. Incubation with insulin like growth factor (IGF) I or IGF-II promoted cell growth, which was reversed by wogonin co-administration. Mechanically, we found that down-regulation of IGF-1R diminished the chemosensitization role of wogonin in breast cancer. In addition, wogonin suppressed the phosphorylation levels of AKT and addition of AKT inhibitor abolished the synergistic cytotoxicity of wogonin and doxorubicin. Taken together, combined treatment with wogonin increased the doxorubicin sensitivity in breast cancer cells through regulation of IGF-1R/AKT signaling pathway. Therefore, these findings demonstrated that combination therapy with wogonin led to better therapeutic effects via regulating IGF-1R/AKT signaling pathway in doxorubicin-based chemotherapy for breast cancer.

Light-activated tissue bonding for excisional wound closure: a split-lesion clinical trial.

BACKGROUND: Apposition of wound edges by sutures provides a temporary scaffold and tension support for healing. We have developed a novel tissue-sealing technology, photoactivated tissue bonding (PTB), which immediately crosslinks proteins between tissue planes, thereby sealing on a molecular scale. OBJECTIVES: To determine the effectiveness of PTB for superficial closure of skin excisions and to compare the results with standard epidermal suturing. METHODS: A split-lesion, paired comparison study of 31 skin excisions was performed. Following deep closure with absorbable sutures, one-half of each wound was superficially closed with nonabsorbable nylon sutures while the other half was stained with Rose Bengal dye and treated with green light. Overall appearance and scar characteristics were rated at 2weeks and 6months in a blinded manner by three dermatologists viewing photographs, by two onsite physicians and by patients. RESULTS: At 2weeks, neither sutured nor PTB-treated segments showed dehiscence; however, PTB-sealed segments showed less erythema than sutured segments as determined by photographic (P=0·001) and onsite evaluations (P=0·005). Overall appearance after PTB was judged better than after sutures (P=0·002). At 6months, scars produced by PTB were deemed superior to scars resulting from sutures in terms of appearance (P<0·001), width (P=0·002) and healing (P=0·003). Patients were more satisfied with the appearance of the PTB-sealed wound half after 2weeks and 6months (P=0·013 and P=0·003, respectively). CONCLUSIONS: A novel molecular suturing technique produces effective wound sealing and less scarring than closure with nylon interrupted epidermal sutures. Comparisons with better suturing techniques are warranted.

In vitro characterization and pharmacokinetics of dapagliflozin (BMS-512148), a potent sodium-glucose cotransporter type II inhibitor, in animals and humans.

(2S,3R,4R,5S,6R)-2-(3-(4-Ethoxybenzyl)-4-chlorophenyl)-6-hydroxymethyl-tetrahydro-2H-pyran-3,4,5-triol (dapagliflozin; BMS-512148) is a potent sodium-glucose cotransporter type II inhibitor in animals and humans and is currently under development for the treatment of type 2 diabetes. The preclinical characterization of dapagliflozin, to allow compound selection and prediction of pharmacological and dispositional behavior in the clinic, involved Caco-2 cell permeability studies, cytochrome P450 (P450) inhibition and induction studies, P450 reaction phenotyping, metabolite identification in hepatocytes, and pharmacokinetics in rats, dogs, and monkeys. Dapagliflozin was found to have good permeability across Caco-2 cell membranes. It was found to be a substrate for P-glycoprotein (P-gp) but not a significant P-gp inhibitor. Dapagliflozin was not found to be an inhibitor or an inducer of human P450 enzymes. The in vitro metabolic profiles of dapagliflozin after incubation with hepatocytes from mice, rats, dogs, monkeys, and humans were qualitatively similar. Rat hepatocyte incubations showed the highest turnover, and dapagliflozin was most stable in human hepatocytes. Prominent in vitro metabolic pathways observed were glucuronidation, hydroxylation, and O-deethylation. Pharmacokinetic parameters for dapagliflozin in preclinical species revealed a compound with adequate oral exposure, clearance, and elimination half-life, consistent with the potential for single daily dosing in humans. The pharmacokinetics in humans after a single dose of 50 mg of [(14)C]dapagliflozin showed good exposure, low clearance, adequate half-life, and no metabolites with significant pharmacological activity or toxicological concern.

Effect of mycorrhization on the accumulation of rishitin and solavetivone in potato plantlets challenged with Rhizoctonia solani.

The effect of colonization with the vesicular-arbuscular mycorrhizal fungus Glomus etunicatum on the content of rishitin and solavetivone was determined in potato plants cv. Goldrush challenged with Rhizoctonia solani. Mycorrhization stimulated significantly the accumulation of both phytoalexins in roots of plantlets challenged with R. solani but did not influence phytoalexin levels in non-challenged plantlet roots. No accumulation of solavetivone or rishitin was detected in shoots. In Petri dish bioassays, rishitin and solavetivone inhibited mycelial growth of R. solani.

Removal and recovery of phosphate and ammonium as struvite from supernatant in anaerobic digestion.

Removal of phosphorus and nitrogen is required to prevent eutrophication problems in lakes and enclosed coastal seas. And recovery of phosphorus from wastewater has been attracting attention because of lack in phosphorus resources in the near future. In this study, reaction kinetics and design parameters of struvite production are experimentally investigated by using basic reaction type and a draft-tube type reactors. Struvite production rate, which is a very important parameter in reactor design and efficiency estimation, is formulated in an equation consisting of a rate constant (k2), and magnesium, phosphate and ammonium concentrations. The value of k2 is shown to be increased with struvite concentration and mixing intensity in the reactor. The developed equation is applied to the results obtained from the draft-tube type reactor experiments and verified for its applicability. High struvite concentration of 10-25% is maintained in the draft-tube reactor experiments. 92% removal and recovery efficiency with effluent phosphorus concentration of 17 mg/L is achieved under the conditions of 4 minutes reaction time, pH of 8.5 and Mg/P molar ratio of 1.1.

Effect of two vesicular-arbuscular mycorrhizal fungi on the growth of micropropagated potato plantlets and on the extent of disease caused by Rhizoctonia solani.

Two micropropagated potato cultivars, Goldrush and LP89221, were inoculated into sowing trays with either Glomus etunicatum or G. intraradices in a greenhouse. After 2 weeks, plantlets were transplanted into pots and roots were challenged 7 days later with Rhizoctonia solani. At different times after R. solani infection, disease severity, mortality rate, root colonization levels, various growth parameters, and shoot mineral content were evaluated. In Goldrush, only inoculation with G. etunicatum led to a significant reduction in disease severity, ranging between 60.2% and 71.2%, on both shoot and crown. This decrease was not observed in LP89221. Compared with the control plantlets, inoculation of Goldrush with G. etunicatum or G. intraradices reduced significantly the mortality rate by 77% and 26%, respectively, whereas vesicular-arbuscular mycorrhizal (VAM) fungi did not significantly influence the mortality rate in LP89221. In Goldrush, inoculation with G. etunicatum significantly increased shoot fresh weight, root dry weight and the number of tubers produced per plant, whereas G. intraradices only significantly increased the number of tubers. Tuber and root fresh weights of both potato cultivars were significantly reduced by R. solani infection. However, R. solani-infected plantlets of both Goldrush and LP89221, inoculated with G. etunicatum, produced significantly greater tuber fresh weight than non-VAM plantlets. In R. solani-infected plantlets of Goldrush but not LP89221, G. etunicatum and G. intraradices increased root fresh weight by approximately 140.3% and 76.5%, respectively, compared with non-VAM plants. The potato cultivars Goldrush and LP89221 responded differently to VAM fungal inoculation and to R. solani infection in terms of shoot mineral content.

Clinical and biological implications of partial tandem duplication of the MLL gene in acute myeloid leukemia without chromosomal abnormalities at 11q23.

The clinical and biological features of acute myeloid leukemia (AML) with 11q23/MLL translocations are well known, but the characteristics of AML with partial tandem duplication of the MLL gene have not been explored comprehensively. In this study, MLL duplication was analyzed, in 81 AML patients without chromosomal abnormalities at 11q23, using Southern blotting, genomic DNA polymerase chain reaction (PCR), reverse-transcription PCR and complementary DNA sequencing. Nine patients showed partial tandem duplication of the MLL gene, including eight (12%) of the 68 with normal karyotype. Seven patients showed fusion of exon 6/exon 2 (e6/e2), one, combination of differentially spliced transcripts e7/e2 and e6/e2, and the remaining one, combination of e8/e2 and e7/e2. Among the patients with normal karyotype, children aged 1 to 15 showed a trend to higher frequency of MLL duplication than other patients (2/5 or 40% vs 6/62 or 10%, P = 0.102). The patients with tandem duplication of the MLL gene had a significantly higher incidence of CD11b expression on leukemic cells than did those without in the subgroup of patients with normal karyotype (75% vs 28%, P = 0.017). There were no significant differences in the expression of lymphoid antigens or other myeloid antigens between the two groups of patients. In adults, the patients with MLL duplication had a shorter median survival time than those without (4.5 months vs 12 months, P = 0.036). In conclusion, partial tandem duplication of the MLL gene is associated with increased expression of CD11b on leukemic blasts and implicates poor prognosis in adult AML patients. The higher frequency of MLL duplication in children older than 1 year, than in other age groups, needs to be confirmed by further studies.

Heart failure and apoptosis: electrophoretic methods support data from micro- and macro-arrays. A critical review of genomics and proteomics.

The multiple causes and multiple consequences of mammalian heart failure make it an attractive proposition for analysis using gene array technology, especially where the failure is idiopathic in nature. However, gene arrays also hold potential artefacts, particularly when gene expression levels are low, and where changes in expression levels are modest. Also, at present, the number of genes available on arrays is not large enough to prevent potential sampling deficiencies. Thus, it may not be wise to place too much reliance on quantitative interpretations of gene array data. Also, recently doubts were raised about the qualitative reliability of array genes. Electrophoretic methods are slow, cumbersome and complex but they can provide confirmation that the trends and numbers arising from the new gene arrays are reliable. In this overview, we compare gene array data with data from protein activity assays such as zymograms, Western blots, two-dimensional electrophoresis, and immunohistochemistry. Similar or complementary data from the same heart tissues analyzed by either microarrays or macroarrays can be reassuring to those interested in reliable molecular analyses of normal and failing hearts. Similar principles will apply to other tissues and cells.

[Effect of tiaoxin recipe on beta-amyloid induced neurotoxic model of the primary cultured rat hippocampal neurons].

OBJECTIVE: To study the effect of Tiaoxin Recipe (TXR) on beta-amyloid induced neurotoxicity in the primary cultured hippocampal neurons and the therapeutical mechanism of it. METHODS: In order to establish neurotoxic model, the primary cultured rat hippocampal neurons were treated with 25 mumol/L aggregated beta-amyloid fragment 1-42 (A beta 1-42). Neuronal survival was assessed by lactate dehydrogenase (LDH) activity. Apoptosis of neurons was determined with transmission electron microscopy (TEM) examination and flowcytometric analysis. The protein expression of apoptosis-associated gene (Bcl-2, Bax, c-Jun) were examined by using immunocytochemical SABC method. RESULTS: In the A beta 1-42 treated neurons, the release of LDH was increased, TEM examination revealed that the cell body became shrunken, the chromatin was compacted or the patches of condensed chromatin lay against the nuclear membrane and the smooth surfaced endoplasmic reticulum was expanded. Flowcytometric analysis displayed the peak of hypodiploid DNA content. The expression of Bcl-2 was lowered, while Bax and c-Jun protein were increased. After treated with TXR, the above-mentioned changes were improved with the exception of c-Jun expression. In the Tacrine treated group, only the neuronal survival was increased, other observed criteria had insignificant change. CONCLUSION: TXR could attenuate the neurotoxic action of A beta 1-42 and improve neuronal survival via suppressing apoptotic process. TXR could inhibit the apoptotic process via regulating the expression of bcl-2 and bax. Tacrine could improve neuronal survival, but its inhibition on neuronal apoptosis was not significant.

A precolumn derivatization high-performance liquid chromatographic method with improved sensitivity and specificity for the determination of astragaloside IV in Radix Astragali.

A reversed-phase high-performance liquid chromatographic method is developed for the determination of astragaloside (AGS) IV, which is known as the active constituent of Radix Astragali. The method uses precolumn derivatization with benzoyl chloride to form the benzoyl ester of AGS IV quantitatively and is carried out with a wide-ranging concentration (0.004-0.080 mg/mL) of the derivatized AGS IV. The eluent consists of 90% methanol, 4% tetrahydrofuran, 6% water, and 0.2% triethylamine, with vitamin D3 added as the internal standard.

Production and crystallization of lobster muscle tropomyosin expressed in Sf9 cells.

A new form of muscle tropomyosin crystal has been obtained, by employing new strategies in protein preparation and crystallization. Non-polymerizable tropomyosin was prepared by removing 11 amino acids at the C-terminus. The truncated tropomyosin was expressed in Sf9 insect cells by use of the baculovirus-based expression system, to obtain highly homogeneous protein preparations. By routinely monitoring homogeneity by mass spectrometry, we found that the homogeneity played a key role in obtaining good crystals. The crystal quality was also dependent on isoforms; the crystals raised from a slow muscle-specific isoform diffracted to a higher resolution, compared with a fast muscle-specific counterpart. For crystallization, a high concentration of organic solvent was used as the precipitant; in the presence of 35% DMSO, tetragonal crystals were formed, which belong to space group P4(3)(1)2(1)2 with cell constants of a=b=105.6 angstrom, c=506.9 angstrom. The crystals gave rise to reflections the intensities of which were characteristically determined by the transform of alpha-helical coiled-coil. Thus in the region of 10-5.5 angstrom resolut along the c*-axis, the reflections were weak. For accurate measurement of these reflection intensities, beam-line ID2 in ESRF Grenoble was advantageous owing to the high brilliance and a low background. There the crystals diffracted to beyond 3.0 A along the c*-axis, whereas along the a*-b*-plane reflections were limited to 6.6 angstrom. Data analysis is under way on a data set from a PtCl4 derivative.

Neuronal differentiation of P19 embryonal carcinoma cells in defined media.

The P19 embryonal carcinoma cell line is a useful model system for analyzing the factors that regulate neuronal differentiation. In order to analyze the extrinsic factors that are involved in differentiation, it is necessary to carry out experiments in fully defined media. Here we have investigated the neuronal differentiation of P19 cells in two defined media. Cells that are propagated and induced with retinoic acid in standard serum-containing medium are capable of differentiating into neuron-like cells in N2 medium. Dividing fibroblast-like cells also appeared in these cultures. After about 10 days in culture in N2 medium, the great majority of neuron-like cells died. On the other hand, culturing induced cells in N2 medium for 5 days and then switching to a defined medium consisting of Neurobasal medium plus B27 supplement allowed the neuron-like cells to survive for prolonged periods of time. This defined medium thus provides a suitable system for analyzing extrinsic factors that affect the survival and differentiation of P19 neurons. P19 cells induced with retinoic acid and plated in N2 were exposed to bFGF and EGF, which are known to be mitogens for neuronal precursor cells. Both growth factors were mitogenic for a subpopulation of the induced cells. In separate experiments, cells cultured in N2 in the presence of RA were induced to differentiate into neuron-like cells.

Alangionosides G-M: glycosides of megastigmane derivatives from the leaves of Alangium premnifolium.

Phytochemical investigation of the 1-BuOH-soluble fraction of the MeOH extract of Alangium premnifolium resulted in the isolation of ten megastigmane glycosides, of which, two, (6R,9R)-3-oxo-alpha-ionol apiofuranosylglucopyranoside and roseoside with (6S,9R)-blumeol A as an aglycone were known. The structures of the eight new compounds were determined by spectroscopic methods.

Embryonic stem cells express neuronal properties in vitro.

Mouse embryonic stem (ES) cells cultured as aggregates and exposed to retinoic acid are induced to express multiple phenotypes normally associated with neurons. A large percentage of treated aggregates produce a rich neuritic outgrowth. Dissociating the induced aggregates with trypsin and plating the cells as a monolayer results in cultures in which a sizable percentage of the cells have a neuronal appearance. These neuron-like cells express class III beta-tubulin and the neurofilament M subunit. Induced cultures express transcripts for neural-associated genes including the neurofilament L subunit, glutamate receptor subunits, the transcription factor Brn-3, and GFAP. Levels of neurofilament L and GAD67 and GAD65 transcripts rise dramatically upon induction. Physiological studies show that the neuron-like cells generate action potentials and express TTX-sensitive sodium channels, as well as voltage-gated potassium channels and calcium channels. We conclude that a complex system of neuronal gene expression can be activated in cultured ES cells. This system should be favorable for investigating some of the mechanisms that regulate neuronal differentiation.

One-megabase yeast artificial chromosome and 400-kilobase cosmid-phage contigs containing the von Hippel-Lindau tumor suppressor and Ca(2+)-transporting adenosine triphosphatase isoform 2 genes.

We have isolated and ordered yeast artificial chromosomes (YACs) and cosmids surrounding the von Hippel-Lindau (VHL) tumor suppressor and plasma membrane Ca(2+)-transporting ATPase isoform 2 (PMCA-2) genes on chromosome 3p25-26. The YAC contig consists of six YACs and covers a region of 1 megabase. A cosmid-phage contig around VHL and PMCA-2 genes (400 kilobases) was established and integrated into the YAC map. Using these clones, we generated an EcoRI map of the 400-kilobase region. PMCA-2 and VHL complementary DNA were positioned entirely within the cosmid-phage contig as well as two polymorphic markers (D3S601 and D3S1317). This physical map of the cloned region will allow a detailed analysis of both the PMCA-2 and VHL genes. Some of the genomic clones may be useful for isolation of the full-length VHL complementary DNA.