An antitumor compound julibroside J28 from Albizia julibrissin

A new triterpenoid saponin, julibroside J(28) (1), was isolated from the stem bark of Albizia julibrissin Durazz (Leguminosae) by using chromatographic method. The structure of 1 was established by spectroscopic methods. 1 displayed significant antitumor activity in vitro against PC-3M-1E8, Bel-7402, and HeLa cancer cell lines at 10 microM assayed by SRB method.     

Two diastereomeric saponins with cytotoxic activity from Albizia julibrissin

Two diastereomeric saponins, julibrosides J1 (1) and J9 (2), both of which show cytotoxic activity, were obtained from the stem bark of Albizia julibrissin Durazz. On the basis of chemical and spectral evidence [L.B. Ma et al., Carbohydr. Res., 281 (1996) 35-46], the structure of 1 was revised as 3-O-[beta-D-xylopyranosyl-(1-->2)-alpha-L-arabinopyranosyl-(1-->6) -beta-D-glucopyranosyl]-21-O-[(6S)-2-trans-2-hydroxymethyl-6-methyl-6-O- [4-O-((6R)-2-trans-2,6-dimethyl-6-O-(beta-D-quinovopyranosyl)-2,7- octadienoyl)-beta-D-quinovopyranosyl]-2,7-octadienoyl] acacic acid-28-O-beta-D-glucopyranosyl-(1-->3)-[alpha-L-arabinofuranosyl-(1-->4 )]-alpha-L-rhamnopyranosyl-(1-->2)-beta-D-glucopyranosyl ester. The diastereoisomer 2 of 1 was identified as 3-O-[beta-D-xylopyranosyl-(1-->2)-alpha-L-arabinopyranosyl-(1-->6) -beta-D-glucopyranosyl]-21-O-[(6S)-2-trans-2-hydroxymethyl-6-methyl-6-O- [4-O-((6S)-2-trans-2,6-dimethyl-6-O-(beta-D-quinovopyranosyl)-2,7- octadienoyl)-beta-D-quinovopyranosyl]-2,7-octadienoyl] acacic acid-28-O-beta-D-glucopyranosyl-(1-->3)-[alpha-L-arabinofuranosyl-(1-->4 )]-alpha-L-rhamnopyranosyl-(1-->2)-beta-D-glucopyranosyl ester. Saponin 2 is a new saponin named julibroside J9. Both julibrosides J1 and J9 show good inhibitory action against the KB cancer cell line in vitro.     

Anti-angiogenic activity of julibroside J8, a natural product isolated from Albizia julibrissin

PurposeThe purpose of this study was to investigate the anti-angiogenic properties of julibroside J₈, a triterpenoid saponin isolated from Albizia julibrissin.MethodsIn the presence of juliborside J_8, the growth of human microvascular endothelial cells (HMEC-1), four human tumor cell lines, and a normal cell line (MRC-5) was evaluated by MTT assay. The in vivo anti-angiogenic effect of julibroside J₈ was evaluated on a chorioallantoic membrane (CAM) and in transplanted colon carcinoma cells in a nude mice neovascularisation model.ResultsTreatment with 0.5–4 μg/ml julibroside J₈ resulted in dose-dependent inhibition of growth, migration, and tube formation in HMEC-1 cells; julibroside J₈ also inhibited the formation of microvessels on CAM at a concentration of 10–50 μg/egg and reduced vessel density within tumor at a concentration of 0.5–3 mg/kg.ConclusionsJulibroside J₈ may be a potent anti-angiogenetic and cytotoxic drug; further investigation is warranted.     

New Cytotoxic Triterpenoid Saponins from the Roots of Albizia gummifera (J.F.Gmel.) C.A.Sm.

As part of our search for new bioactive saponins from Cameroonian medicinal plants, two new oleanane‐type saponins, named gummiferaosides D and E ( 1 and 2 ), along with one known saponin, julibroside J₈ ( 3 ), were isolated from the roots of Albizia gummifera. Their structures were established on the basis of extensive 1D‐ and 2D‐NMR (¹H‐ and ¹³C‐NMR, DEPT, COSY, TOCSY, NOESY, HSQC, HSQC‐TOCSY, and HMBC) and HR‐ESI‐MS studies, and by chemical evidence. The apoptotic effect of saponins 1  –  3 was evaluated on the A431 human epidermoid cancer cell. Flow cytometric analyses showed that saponins 1  –  3 induced apoptosis of human epidermoid cancer cell (A431) in a dose‐dependent manner.     

黔产青钱柳化学成分及α-葡萄糖苷酶抑制活性研究

为了阐明青钱柳中的降血糖功效成分,实验对黔产青钱柳进行了系统化学成分研究和α-葡萄糖苷酶抑制活性评价。利用各种色谱学手段和波谱学方法,从青钱柳叶80%乙醇提取物中分离鉴定出13个化合物,分别为阿福豆苷(1)、山柰酚3-O-(4″-O-乙酰基)-α-L-吡喃鼠李糖苷(2)、juglanoside J(3)、1α,2α,4β-3羟基-1,2,3,4-四氢萘酮(4)、青钱柳苷III(5)、青钱柳苷J(6)、黄体酮(7)、去酰基萝藦苷元(8)、20-乙酰氧基-4-烯-3-酮(9)、齐墩果酸(10)、5-O-P香豆酰奎宁酸甲酯(11)、亚麻酸甲酯(12)、软脂酸(13)。其中,化合物9为新天然产物,化合物2~4、7、8、11、13均为首次从该植物中分离得到。用PNPG法对化合物1~10进行了α-葡萄糖苷酶抑制活性筛选,结果显示,化合物1、2有较强的α-葡萄糖苷酶抑制活性,表明黄酮类成分是青钱柳降血糖功效的物质基础之一。     

A four-way junction with triple-helical arms: design, characterization, and stability

The formation of the four-way junction containing four triple-helical arms has been demonstrated using chemical methods (polyacrylamide gel electrophoresis and chemical footprinting using OsO(4) as a probe) and physical methods (UV absorbance melting and DSC). The junction J(T1T3) was assembled from two 20-mer purine strands and two 44-mer pyrimidine strands. To determine the contribution of the different arms to the stability of the complete structure of J(T1T3), the junction was compared to two simplified substructures, J(T1) and J(T3), respectively. Common to these complexes is the underlying double-helical four-way junction Js. Addition of Na(+) had a profound effect on stabilizing and subsequently folding the junctions into the stacked X-structures. The following results support the structure present: (i) The native polyacrylamide electrophoresis exhibits only a single band(s) corresponding to one species present when all four single strands are mixed in equal amounts. (ii) OsO(4) modifications were investigated at pH 5.0 and in the presence of 10 mM Mg(2+) and 100 mM Na(+). There is no cleavage of thymine residues at the branch point and throughout the structure. (iii) The thermal unfolding of J(T1) and J(T3) illustrates that the triple-helical arms are more stable than the double-helical arms which are contained in these junctions and that J(T1T3) with four triple-helical arms is slightly more stable than J(T1) and J(T3). (iv) The calorimetric transition enthalpies determined for the arms of J(T1T3) are comparable to those associated with the unfolding of its corresponding arms in J(T1) and J(T3). The results also illustrate that the formation of the junctions is not restricted by the pH, [Na(+)], sequence composition of the arms, and/or the loop position.     

Two γ-Methyl Biosides from Dregea volubilis (L.) Benth

From the hydrolysate of the crude glycosides from the roots, of Dregea volubilis(L.) Benth in Dehong, Yunnan, two α-methyl biosides Ⅰ and Ⅱ (yields: 0.016%and 0.0097%, respectively) were isolated by silica gel column chromatography. Theirchemical structures were established by interpretation of MS, IR,1H,13C-NMR, andgas chromatographic analysis of their degradation products, and comparison of thephysical properties of Ⅰ, Ⅱ and their acetates which were reported in literatures asfollows: α-methyl-pachybioside for Ⅰ, and α-methyl-[3-O-methyl-6-deoxy-D-allose(1→4)-D-olivoside] for Ⅱ. Ⅱ named α-methyl-dredehongbioside, is reported for the first time. Ⅱ, α-methyl-dredehongbioside, colorless needles (from MeOH), bitter, mp. 184–186℃,[α]D22 +74.5˚~(c= 0.52, MeOH). Anal. Cald(%) for C14H26O8:C52.17, H8.07;Found; C52.23,H8.22. Irvmaxkbr: 3370, 1443, 1419, 1375, 1268, 1218, 1168,1127,1060cm-1. MS(m/e,%): 322(M+,3),291(M+-OCH3,15), 273(M+-OCH3-H2O,12), 258,246,232, 222, 159, 145, 141, 128, 95, 87, 85, 74 (base peak, 100), 59. 1H NMRδ(CDCl3): 4.73(1H, dd, J= 4.0 Hz, J= 1.5Hz, C-1-H), 4.55(1H, d, J= 8.0Hz,C-1′-H), 3.79(1H, dd, J=3.0Hz, J= 3.0Hz, C-3′-H), 3.00(1H, dd, J= 9.0Hz,J= 9.0Hz, C-4-H), 2.22(1H, m, C-2-Ha), 1.60(1H, m, C-2-He), 1.33(3H, d,J= 6.0Hz,C-5-CH,), 1.31(3H,d,,J= 6.5Hz, C-5′-CH), 3.68(3H,s,,C-3′-OCH),3.31(3H,s,C-1-OCH). 13C NMR data were seen in Table 1. Ⅳ, tri-acetyl-α-methyl-dredehongbioside, colorless granular (from MeOH),mp. 135--137℃, [a]D22+ 88.2˚(c= 0.50, MeOH). MS(m/e, %): 488 (M+, 2), 388(M+-HOAc,2), 357(M+-OCH3-HOAc,33), 288, 187, 127, 116, 85, 74, 59, 43(basepeak, 100). 1H NMR,δ(CDCl3): 5.25(1H, ddd,.J= 11.0Hz, J=9.0Hz,.J= 5.5Hz,C-3-H), 4.86(1H, d, J= 8.0Hz, C-1′-H), 4.69 (1H, dd, J= 4.0Hz, J= 1.5Hz,C-1-H), 4.58(1H,m,C-2′-H),3.94(1H, dd, J= 3.0Hz,J= 3.0Hz,C-3′-H),3.64(1H,m,C-5-H), 3.22(1H,dd, J= 9.5Hz, J= 8.5Hz, C-4-H), 2.30(1H, m, C-2-Ha),1.67(1H,m,C-2-He), 1.31(3H, d, .J= 6.5Hz, C-5-CH3), 1.17(3H, d, J= 6.0Hz,C-5-CH3), 3.47(3H, s, C-3′-OCH3), 3.30(3H,s, C-1-OCH3), 2.10(6H, s, C-2′, C- 4′ -OCH3), 2.03(3H,s,C-3-OCH3).     

H2 control of natural T regulatory cell frequency in the lymph node correlates with susceptibility to day 3 thymectomy-induced autoimmune disease

Day 3 thymectomy (D3Tx) results in a loss of peripheral tolerance mediated by natural regulatory T cells (nTregs) and development of autoimmune ovarian dysgenesis (AOD) and autoimmune dacryoadenitis (ADA) in A/J and (C57BL/6J × A/J) F(1) hybrids (B6A), but not in C57BL/6J (B6) mice. Previously, using quantitative trait locus (QTL) linkage analysis, we showed that D3Tx-AOD is controlled by five unlinked QTL (Aod1-Aod5) and H2. In this study, using D3Tx B6-Chr(A/J)/NaJ chromosome (Chr) substitution strains, we confirm that QTL on Chr16 (Aod1a/Aod1b), Chr3 (Aod2), Chr1 (Aod3), Chr2 (Aod4), Chr7 (Aod5), and Chr17 (H2) control D3Tx-AOD susceptibility. In addition, we also present data mapping QTL controlling D3Tx-ADA to Chr17 (Ada1/H2), Chr1 (Ada2), and Chr3 (Ada3). Importantly, B6-ChrX(A/J) mice were as resistant to D3Tx-AOD and D3Tx-ADA as B6 mice, thereby excluding Foxp3 as a susceptibility gene in these models. Moreover, we report quantitative differences in the frequency of nTregs in the lymph nodes (LNs), but not spleen or thymus, of AOD/ADA-resistant B6 and AOD/ADA-susceptible A/J, B6A, and B6-Chr17(A/J) mice. Similar results correlating with experimental allergic encephalomyelitis and orchitis susceptibility were seen with B10.S and SJL/J mice. Using H2-congenic mice, we show that the observed difference in frequency of LN nTregs is controlled by Ada1/H2. These data support the existence of an LN-specific, H2-controlled mechanism regulating the prevalence of nTregs in autoimmune disease susceptibility.     

New Karplus equations for 2JHH, 3JHH, 2JCH, 3JCH, 3JCOCH, 3JCSCH, and 3JCCCH in some aldohexopyranoside derivatives as determined using NMR spectroscopy and density functional theory calculations

The (1)H-(13)C coupling constants of methyl alpha- and beta-pyranosides of D-glucose and D-galactose have been measured by one-dimensional and two-dimensional (1)H-(13)C heteronuclear zero and double quantum, phase sensitive J-HMBC spectra to determine a complete set of coupling constants ((1)J(CH), (2)J(CH), (3)J(CH), (2)J(HH), and (3)J(HH)) within the exocyclic hydroxymethyl group (CH(2)OH) for each compound. In parallel with these experimental studies, structure, energy, and potential energy surfaces of the hydroxymethyl group for these compounds were determined employing quantum mechanical calculations at the B3LYP level using the 6-311++G( * *) basis set. Values of the vicinal coupling constants involving (1)H and (13)C in the C5-C6 (omega) and C6-O6 (theta) torsion angles in the aldohexopyranoside model compounds were calculated with water as the solvent using the PCM method. To test the relationship between (3)J(CXCH) (X=C, O, S) and torsion angle C1-X (phi) around the anomeric center, the conformations of 24 derivatives of glucose and galactose, which represent sequences of atoms at the anomeric center of C-glycosides (C-C bond), O-glycosides (C-O bond), thioglycosides (C-S bond), glycosylamines (C-N bond), and glycosyl halides (C-halogen (F/Cl) bond) have been calculated. Nonlinear regression analysis of the coupling constants (1)J(C1,H1), (2)J(C5,H6R), (2)J(C5,H6S), (2)J(C6,H5), (3)J(C4,H6R), (3)J(C4,H6S), (2)J(H6R,H5), and (3)J(H5,H6R) as well as (3)J(CXCH) (X=C, O, S) on the dihedral angles omega, theta, and phi have yielded new Karplus equations. Good agreement between calculated and experimentally measured coupling constants revealed that the DFT method was able to accurately predict J-couplings in aqueous solutions.     

菌根真菌与施肥对杜鹃苗养分吸收的影响

以2年生杜鹃(Rhododendron simsii)苗为研究对象,采用J1(杜鹃花类菌根真菌混合接菌)、J2(杜鹃花类菌根真菌和马尾松外生菌混合接菌)、J3(不接菌,对照)等3种接菌组合与不同氮磷钾施肥组合研究菌根真菌与氮磷钾肥对杜鹃苗生长及养分吸收的影响。结果表明,杜鹃幼苗的生长指标和氮磷钾含量总体表现为接菌处理优于不接菌处理,J2处理下施用尿素1.1 g·株-1、钙镁磷1 g·株-1、氯化钾0.7 g·株-1对杜鹃苗生长及氮磷钾含量积累的促进作用最为明显。数据表明,J2处理促进N向叶积累,促进K向茎叶积累,促进P向根茎积累;J1处理促进N和P向根茎积累,促进K向茎叶积累。     

褐薄小齿菌的化学成分研究

从褐薄小齿菌(Hydnellum concrescens)子实体中分离得到9个已知化合物,借助光谱手段,它们的化学结构分别鉴定为:friedelin(1),(22E,24R)-麦角甾-5,7,22-三烯-3β-醇(2),5α,8α-过氧-(22E,24R)-麦角甾-6,22-二烯-3β-醇(3),(22E,24R)-麦角甾-5,22-二烯-3β-羟基-7-酮(4),6-甲氧基-cerevisterol(5),thelephantin K(6),thelephantin I(7),thelephantin J(8),thelephantin L(9)。     

The formation of the oncofetal J28 glycotope involves core-2 beta6-N-acetylglucosaminyltransferase and alpha3/4-fucosyltransferase activities.

The feto-acinar pancreatic protein or FAPP, the oncofetal glycoisoform of bile salt-dependent lipase (BSDL), is characterized by the presence of the J28 glycotope recognized by mAbJ28. This fucosylated epitope is carried out by the O-linked glycans of the C-terminal mucin-like region of BSDL. This glycotope is expressed by human tumoral pancreatic tissues and by human pancreatic tumoral cell lines such as SOJ-6 and BxPC-3 cells. However, it is not expressed by the normal human pancreatic tissues and by MiaPaCa-2 and Panc-1 cells. Due to the presence of many putative sites for O-glycosylation on FAPP and BSDL, the structure of the J28 glycotope cannot be attained by classical physical methods. In the first part of the present study, we have determined which glycosyltransferases were differently expressed in pancreatic tumoral cell lines compared to normal tissues, focusing in part on fucosyltransferases (Fuc-T) and core-2 beta6-N-acetylglucosaminyltransferase (Core2GlcNAc-T). Our data suggested that alpha2-Fuc-T activity was decreased in the four cell lines tested (SOJ-6, BxPC-3, MiaPaCa-2, and Panc-1). The alpha(1-3) and alpha(1-4) fucosylations were decreased in tumor cells that do not express the J28 glycotope whereas alpha4-Fuc-T and Core2GlcNAc-T activities were significantly increased in SOJ-6 cells which best expressed the J28 glycotope. Therefore, we wished to gain information about glycosyltransferases involved in the building of this structure by transfecting the cDNA encoding the mucin-like region of BSDL in CHO-K1 also expressing Core2GlcNAc-T and/or FUT3 and/or FUT7 activities. These CHO-K1 cells have been previously transfected with the cDNA encoding Core2GlcNAc-T and/or FUT3 and/or FUT7. Data indicated that the C-terminal peptide of BSDL (Cter) produced by those cells did not carry out the J28 glycotope unless Core2GlcNAc-T activity is present. Further transfection with FUT3 cDNA, increased the antibody recognition. Nevertheless, transfection with FUT3 or FUT7 alone did not generate the formation of the J28 glycotope on the C-terminal peptide. Furthermore, the Cter peptide produced by CHO-K1 cells expressing Core2GlcNAc-T was more reactive to the mAbJ28 after in vitro fucosylation with the recombinant soluble form of FUT3. These data suggested that the J28 glycotope encompasses structures initiated by Core2GlcNAc-T and further fucosylated by alpha3/4-Fuc-T such as FUT3, likely on GlcNAc residues.     

An endoplasmic reticulum (ER) stress-suppressive compound and its analogues from the mushroom Hericium erinaceum

Three new compounds, 3-hydroxyhericenone F (1), hericenone I (2), and hericenone J (3), were isolated from the mushroom Hericium erinaceum. The structures of 1-3 were determined by the interpretation of spectral data. Compound 1 showed the protective activity against endoplasmic reticulum (ER) stress-dependent Neuro2a cell death, however, compounds 2 and 3 did not.     

Corneal and stomach expression of aldehyde dehydrogenases: from fish to mammals

We have studied the distribution of the ALDH3A1, ALDH1A1 and ALDH2 proteins in the cornea and stomach of several animal species, including mammals (C57BL/6J and SWR/J mice, rat and pig), birds (chicken and turkey), amphibians (frog) and fish (trout and zebrafish). High ALDH3A1 protein levels and catalytic activities were detected in C57BL/6J mouse, rat and pig. We found complete absence of the ALDH3A1 protein in SWR/J mice, which carry the Aldh3a1(c) allele characterized by four amino acid substitutions (G88R, I154N, H305R and I352V) and lack of enzymatic activity. This indicates that the SWR/J mouse strain is a natural gene knockout model for ALDH3A1. Traces of ALDH3A1 were detected in rabbit, whereas expression was absent from chicken, turkey, frog, trout, and zebrafish. Interestingly, significant levels of the cytosolic ALDH1A1 and mitochondrial ALDH2 proteins were detected by immunoblot analysis in all examined species that are deficient in ALDH3A1 expression. In contrast, no ALDH1A1 or ALDH2 protein was detected in the species expressing ALDH3A1. It can, therefore, be concluded that corneal expression of ALDH3A1 or ALDH1A1/ALDH2 occurs in a taxon-specific manner, supporting the protective role of these ALDHs in cornea against the UV-induced oxidative damage.     

Regulation of glomerulotubular balance. I. Impact of dopamine on flow-dependent transport

In response to volume expansion, locally generated dopamine decreases proximal tubule reabsorption by reducing both Na/H-exchanger 3 (NHE3) and Na-K-ATPase activity. We have previously demonstrated that mouse proximal tubules in vitro respond to changes in luminal flow with proportional changes in Na(+) and HCO(3)(-) reabsorption and have suggested that this observation underlies glomerulotubular balance. In the present work, we investigate the impact of dopamine on the sensitivity of reabsorptive fluxes to changes in luminal flow. Mouse proximal tubules were microperfused in vitro at low and high flow rates, and volume and HCO(3)(-) reabsorption (J(v) and J(HCO3)) were measured, while Na(+) and Cl(-) reabsorption (J(Na) and J(Cl)) were estimated. Raising luminal flow increased J(v), J(Na), and J(HCO3) but did not change J(Cl). Luminal dopamine did not change J(v), J(Na), and J(HCO3) at low flow rates but completely abolished the increments of Na(+) absorption by flow and partially inhibited the flow-stimulated HCO(3)(-) absorption. The remaining flow-stimulated HCO(3)(-) absorption was completely abolished by bafilomycin. The DA1 receptor blocker SCH23390 and the PKA inhibitor H89 blocked the effect of exogenous dopamine and produced a two to threefold increase in the sensitivity of proximal Na(+) reabsorption to luminal flow rate. Under the variety of perfusion conditions, changes in cell volume were small and did not always parallel changes in Na(+) transport. We conclude that 1) dopamine inhibits flow-stimulated NHE3 activity by activation of the DA1 receptor via a PKA-mediated mechanism; 2) dopamine has no effect on flow-stimulated H-ATPase activity; 3) there is no evidence of flow stimulation of Cl(-) reabsorption; and 4) the impact of dopamine is a coordinated modulation of both luminal and peritubular Na(+) transporters.     

Inactivation of single-celled Ascaris suum eggs by low-pressure UV radiation

Intact and decorticated single-celled Ascaris suum eggs were exposed to UV radiation from low-pressure, germicidal lamps at fluences (doses) ranging from 0 to 8,000 J/m2 for intact eggs and from 0 to 500 J/m2 for decorticated eggs. With a UV fluence of 500 J/m2, 0.44-+/-0.20-log inactivation (mean+/-95% confidence interval) (63.7%) of intact eggs was observed, while a fluence of 4,000 J/m2 resulted in 2.23-+/-0.49-log inactivation (99.4%). (The maximum quantifiable inactivation was 2.5 log units.) Thus, according to the methods used here, Ascaris eggs are the most UV-resistant water-related pathogen identified to date. For the range of fluences recommended for disinfecting drinking water and wastewater (200 to 2,000 J/m2), from 0- to 1.5-log inactivation can be expected, although at typical fluences (less than 1,000 J/m2), the inactivation may be less than 1 log. When the eggs were decorticated (the outer egg shell layers were removed with sodium hypochlorite, leaving only the lipoprotein ascaroside layer) before exposure to UV, 1.80-+/-0.32-log reduction (98.4%) was achieved with a fluence of 500 J/m2, suggesting that the outer eggshell layers protected A. suum eggs from inactivation by UV radiation. This protection may have been due to UV absorption by proteins in the outer layers of the 3- to 4-microm-thick eggshell. Stirring alone (without UV exposure) also inactivated some of the Ascaris eggs (approximately 20% after 75 min), which complicated determination of the inactivation caused by UV radiation alone.     

Characterization of Carbohydrates on the Surface of Second-stage Juveniles of Meloidogyne spp.

Fluorescent conjugates of the lectins soybean agglutinin (SBA), Concanavalin A (Con A), wheat germ agglutinin (WGA), Lotus tetragonolobus agglutinin (LOT), and Limulus polyphemus agglutinin (LPA) bound primarily to amphidial openings and amphidial secretions of viable, preinfective second-stage juveniles (J2) of Meloidogyne incognita races 1 and 3 (Mil, Mi3) and M. javanica (Mj). No substantial difference in fluorescent lectin binding was observed among the populations examined. Binding of only LOT and LPA were inhibited in the presence of 0.1 M competitive sugar. Structural differences in amphidial carbohydrate complexes among populations of Mi 1, Mi3, and Mj were revealed by glycohydrolase treatment of preinfective J2 and subsequent labeling with fluorescent lectins. A quantitative microfiltration enzyme-linked lectin assay revealed previously undetected differences in lectin binding to nonglycohydrolase-treated J2. Freinfective J2 of Mj bound the greatest amount of SBA, LOT, and WGA, whereas J2 of Mil bound the most LPA.     

Studies on the Alkaloids of Cephalotaxus Isolation of 7-deoxycephalofortuneine,a New Homoerythrina Alkaloid from Cephalotaxus fortuneine Hook. f

Six homoerythrina alkaloids were isolated and identified from the weak base fraction of alkaloids from Cephalotaxus fortunei Hook. f. grown in Anhui Province, China. Five of them were known compounds and identified as cephalofortuneine (2), comosimine (Phelline alkaloid 6) (3), 3-epifortuneine (4), wilsonine (6) and epiwilsonine (7), respectively. One of them is a new alkaloid and proved to be 7-deoxycephalofortuneine (1) on the base of spectral analysis. Its MS shows a molecular ion at m/z 345, 16 mass units less than that of cephalofortuneine (2). Comparison of its 1H-NMR spectrum with that of 2 indicats that 1 lacks 7-hydroxyl group, but it still has 2-hydroxyl group.J3,4ax and J3,4eq are ll.2 Hz and 3.0 Hz, so that 3-Hpresumably is axial. The J1,2 and J2,3 are found to be 4 Hz. These indicat that the 2-H should be equatorial, since J2,3= 4 Hz is small for the trans-axial coupling.     

Fibroblasts that proliferate near denervated synaptic sites in skeletal muscle synthesize the adhesive molecules tenascin(J1), N-CAM, fibronectin, and a heparan sulfate proteoglycan

Four adhesive molecules, tenascin(J1), N-CAM, fibronectin, and a heparan sulfate proteoglycan, accumulate in interstitial spaces near synaptic sites after denervation of rat skeletal muscle (Sanes, J. R., M. Schachner, and J. Covault. 1986. J. Cell Biol. 102:420-431). We have now asked which cells synthesize these molecules, and how this synthesis is regulated. Electron microscopy revealed that mononucleated cells selectively accumulate in perisynaptic interstitial spaces beginning 2 d after denervation. These cells were identified as fibroblasts by ultrastructural and immunohistochemical criteria; [3H]thymidine autoradiography revealed that their accumulation results from local proliferation. Electron microscopic immunohistochemistry demonstrated that N-CAM is associated with the surface of the fibroblasts, while tenascin(J1) is associated with collagen fibers that abut fibroblasts. Using immunofluorescence and immunoprecipitation methods, we found that fibroblasts isolated from perisynaptic regions of denervated muscle synthesize N-CAM, tenascin(J1), fibronectin, and a heparan sulfate proteoglycan in vitro. Thus, fibroblasts that selectively proliferate in interstitial spaces near synaptic sites are likely to be the cellular source of the interstitial deposits of adhesive molecules in denervated muscle. To elucidate factors that might regulate the accumulation of these molecules in vivo, we analyzed the expression of tenascin(J1) and fibronectin by cultured fibroblasts. Fibroblasts from synapse-free regions of denervated muscle, as well as skin, lung, and 3T3 fibroblasts accumulate high levels of tenascin(J1) and fibronectin in culture, showing that perisynaptic fibroblasts are not unique in this regard. However, when they are first placed in culture, fibroblasts from denervated muscle bear more tenascin(J1) than fibroblasts from innervated muscle, indicating that expression of this molecule by fibroblasts is regulated by the muscle's state of innervation; this difference is no longer apparent after a few days in culture. In 3T3 cells, accumulation of tenascin(J1) is high in proliferating cultures, depressed in confluent cultures, and reactivated in cells stimulated to proliferate by replating at low density or by wounding a confluent monolayer. Thus, synthesis of tenascin(J1) is regulated in parallel with mitotic activity. In contrast, levels of fibronectin, which increase less dramatically after denervation in vivo, are similar in fibroblasts from innervated and denervated muscle and in proliferating and quiescent 3T3 cells.(ABSTRACT TRUNCATED AT 400 WORDS)