L), untested as a polydentate ligand, has been found to form the monomeric title complex, [Ni(C12H8N4S)2(H2O)2](BF4)2. The complex shows an octahedral environment of the nickel cation in which the Ni2+ ion is located on a center of symmetry, linked to two ligands and two water molecules. In this 1:2 complex  each thiadiazole ligand uses one pyridyl and one thiadiazole N atom for chelate binding. In the second pyridyl substituent, the N atom is oriented towards the same direction as the S atom of the 1,3,4-thiadiazole ring. The mean plane of the thiadiazole and pyridyl rings linked to the nickel cation forms a dihedral angle with the other pyridine ring of 18.63 (8)°. The tetrafluoridoborate ions can be regarded as free anions in the crystal lattice. Nevertheless, they are involved in an infinite two-dimensional network parallel to -1,3,4-thiadiazole (denoted  II and CuII complexes containing a five azide ring, see: Keij et al. 2(H2O)2](BF4)2                         = 0.029                           wR(F                           2) = 0.073                           S = 1.05                           3120 reflections214 parametersH-atom parameters constrainedmax = 0.60 e Å−3                        Δρmin = −0.40 e Å−3                        Δρ                                 APEX2  used to solve structure: SHELXS97  I, global. DOI: 10.1107/S1600536811026420/dn2703Isup2.hkl            Structure factors: contains datablock(s) I. DOI:  crystallographic information; 3D view; checkCIF report            Additional supplementary materials:


II atoms in the title salt, [Co(H2O)6][Co(C4H4O4S)2]·4H2O, exist in an octahedral coordination environment. In the cation, the Co atom is surrounded by six water molecules, and in the anion, it is bis-O,S,O′-chelated by the thioacetate ligands. The cations, anions and uncoordinated water molecules are linked by O—H⋯O hydrogen bonds into a three-dimensional network.The two Co DOI: 10.1107/S1600536811040979/bt5666Isup2.hkl            Structure factors: contains datablock(s) I. DOI:  crystallographic information; 3D view; checkCIF report            Additional supplementary materials:


In each molecule, the SnIV atom exists within a distorted octahedral geometry defined by the N,N′,S-tridentate mono-deprotonated Schiff base ligand, two mutually trans Cl atoms, and the α-C atom of the n-butyl group; the latter is trans to the azo-N atom. The greatest distortion from the ideal geometry is found in the nominally trans angle formed by the S and pyridyl-N atoms at Sn . In the crystal, molecules are consolidated into a three-dimensional architecture by a combination of N—H⋯Cl, C—H⋯π and π–π interactions [inter-centroid distances = 3.6718 (19) and 3.675 (2) Å].Two independent molecules comprise the asymmetric unit in the title compound, [Sn(C DOI: 10.1107/S1600536812025937/qm2072Isup2.hklStructure factors: contains datablock(s) I. DOI:  crystallographic information; 3D view; checkCIF reportAdditional supplementary materials:


One of the water molecules lies on an inversion centre, so that its H atoms are disordered over two sites. The coordination environment of NiII has a slightly distorted octahedral geometry, which is formed by one O and five N atoms belonging to the N,O-chelating pyrazol-1-ide-5-carboxylate and two N,N′-chelating phenanthroline molecules. In the crystal, O—H⋯O, N—H⋯O and O—H⋯N hydrogen bonds involving the solvent water molecules and pyrazole-5-carboxylate ligands form layers parallel to the ab plane. These layers are linked further via weak π–π interactions between two adjacent phenanthroline molecules, with centroid-to-centroid distances in the range 3.886 (2)–4.018 (1) Å, together with C—H⋯π contacts, forming a three-dimensional network.The title compound, [Ni(C DOI: 10.1107/S1600536813017194/sj5334Isup2.hklStructure factors: contains datablock(s) I. DOI:  crystallographic information; 3D view; checkCIF reportAdditional supplementary materials:


AbstractOmalodes are described from South America, including descriptions of male and female genitalia. Omalodes  mazurisp. n. is described from Peru and Bolivia; Omalodes  punctulatussp. n. from Brazil, and Omalodes  rivussp. n. from Bolivia and Brazil. This is the first paper with detailed information on the male and female genitalia of the genus, showing a great degree of variation to characterize different species. All three new species can be easily distinguished from other known Omalodes and from each other, on the basis of morphology and male genitalia pattern.Three new species of  PageBreakOmalodes was first proposed in the The name Omalodini, the genus had 60 species, divided into 3 subgenera, with Omalodessensu stricto beingthe largest, with 51 species. Given the fact that Omalodes represent close to 70% of the entire tribe . The genus has a neotropical distribution, with only three species being registered for the southern border of the United States, mostly Florida and Arizona, and not being found on the southern half of Argentina and Chile . Each line on the same label is separated by a . Each label is separated by a slash between spaces (/).The photographs were taken with a Leica DFC500 digital camera, edited using the software Auto-montage PRO Digital imaging system (SYNCROSCOPY) and Leica MZ16 stereomicroscope.Museu de Zoologia da Universidade de São Paulo, São Paulo (MZSP), Museu Nacional do Rio de Janeiro, Rio de Janeiro (MNRJ), PageBreakParaná Coleção Entomológica Padre Jesus Santiago Moure, Curitiba, (DZUP), Museu Anchieta, Porto Alegre, Rio Grande do Sul (MAPA), Museu Paraense Emilio Goeldi, Belém, Pará (MPEG), Brazil; Snow Entomological Collection, University of Kansas Natural History Museum, Lawrence, Kansas (SMEK) and Field Museum of Natural History, Chicago, Illinois (FMNH), United States of America. The type material was deposited in the museum from which it was originally borrowed.The studied material belongs to the following institutions: http://zoobank.org/B4C51AD8-179C-4DC0-B44E-FCDD2CD882F4http://species-id.net/wiki/Omalodes_mazuriHolotype: Male. BOLIVIA: “Bolivia tropical; Region Chapare; I:5:50, 400m; R. Zischka leg. / regarded by Mazur as n. sp.” (FMNH). Paratype: Male. PERU: “PERU: Cusco, Villa Carmen Field; Stn., ~1.7 km W cafeteria, res.; trns. 12.89250°S, 71.41917°W; 555m. 22–24.v.2011. Flight; intercept. D. J. Bennett & E. Razuri; PER-11-FIT-007” (SMEK).Frons with a sulcus, little wider close to the epistoma ; suturalPageBreakPageBreakSize range: Length: 5–6.5mm, Width: 4–5mm. Body form  fortunatus Lewis, 1898 has a similar stria. The sutural stria is longer than the usual for the subgenus, being indicated on the posterior half as a well marked, continuous stria. The only species that have the sutural stria as long as this species are Omalodes  pulvintatus Erichson, 1834 and Omalodes  mendax Marseul, 1861, but both have the stria indicated by a series of punctures. In other species of the subgenus the stria is absent or only indicated by a few punctures close to the posterior margin of the elytra. Even though those characters are a little different from usual for the subgenus, this species can be easily placed inside Omalodess. str. when a wider set of characters is taken into account.The complete marginal mesosternal stria of this species is somewhat peculiar in regards to the subgenus The paratype of this species lacks the pair of pronotal foveae close to the lateral punctuation. This probably means that the character is somewhat variable or more probably that those fovea are a developmental anomaly.http://zoobank.org/2BCE1143-798F-4569-81BA-0D7956B43D18http://species-id.net/wiki/Omalodes_rivusHolotype: Male. BRAZIL:“Brasil Pará; Serra Norte; Manganes M2; 13/X/1986 / Armadilha; Shanon / Brasil Pará; J. Dias” (MPEG). Paratypes: BOLIVIA:“Bolívia; Pcia Ichilo; Buenavista XI.1950; A. Martinez leg.”, 1 specimen; (MZSP); BRAZIL: “Brasil, Amapá; Serra do Navio; X-1996; P. Magno leg. / Assentamento”, 1 specimen (MNRJ); “Coleção; M. Alvarenga / Colônia Rio Branco; Óbidos Pará BRASIL; 5-XII-1952; P. A. Teles col.”, 1 specimen (DZUP); “Coleção Campos Seabra / Mangabeira MOCAJUBA; Pará BRASIL; II-1953; Orlando M. Rego”, 1 specimen (MNRJ); “Coleção Campos Seabra / Óbidos; Pará BRASIL; I-1954 J. Brazilino”, 2 specimens (MNRJ); “Brasil; Santaremsinho; Mun. de Itaituba; Rio Tapajós = Pará; VI-62 Dirings”, 1 specimen (MZSP); “Itaituba, PA; XII.1963”, 1 specimen (MZSP); “Brasil, Pará; Marajó-Breves; Rio Caruacá; 22.II.1988 / Area 1; Margem–SE; Isca 1 Solo”, 5 specimens (MPEG); “Brasil, Pará; Marajó-Breves; Rio Caruacá; 22.II.1988 / Area 1; Isca 2; Coleta solo”, 1 specimen (MPEG); “Brasil, Pará; Marajó-Breves; Rio Caruacá; 22.II.1988 / Area 2; Margem W; Isca 2”, 5 specimens (MPEG); “Brasil, Pará; Marajó-Breves; Rio Caruacá; 22.II.1988 / Area 2; Isca 2; Margem W; Coletas solo”, 1 specimen (MPEG); “Brasil Pará; Marajó-Breves; Rio Caruacá; 4.VIII.1988 / Brasil PA; J. Dias / B55; L17”, 1 specimen (MPEG); “Brasil Pará; Marajó-Breves; Rio Caruacá; 6.VIII.1988 / Isca Laranja / M. Martins”, 1 specimen (MPEG); “Brasil Pará; Marajó-Breves; Rio Caruacá; B60; 6.VIII.1988 / Brasil Pará; J. Dias”, 1 specimen (MPEG); “Brasil Pará; Marajó-Breves; Rio Caruacá; 9.VIII.1988; B-66–Isca Laranja / Brasil Pará; J. Dias”, 2 specimens (MPEG); “Brasil Pará; Marajó-Breves; Rio Caruacá; 12.VIII.1988; B72; Isca abacaxi / Brasil Pará; J. Dias”, 1 specimen (MPEG); “Brasil Pará; B72; Marajó-Breves; Rio Caruacá; 12.VIII.1988; Isca laranja / Brasil Pará; J. Dias”, 1 specimen (MPEG); “Brasil Pará; Serra Norte; Pojuca; 4.VII.1985 / Brasil Pará; R. B. Neto”, 1 specimen (MPEG); “Brasil Pará; Serra Norte; M2 Mata; Isca de fruta; 24 a 26.I.1986 / Brasil Pará; M. F. Torres”, 1 specimen (MPEG); “Brasil Pará; Serra Norte; Manganês M2; 13.X.1986 / Armadilha; Shanon / Brasil Pará; J. Dias”, 1 specimens (MPEG); “Brasil Pará; Serra Norte; Manganês; Isca 2.4; 13.X.1986 / Brasil Pará; J. Dias”, 1 specimen (MPEG); “Brasil Pará; Serra Norte; 3 Alfa; 15 a 18.X.1986 / Armadilha Shanon; J. Dias”, 1 specimen (MPEG); PageBreakPageBreak“Brasil Pará; Serra Norte; N2; Isca fruta; 22 a 24.I.1986 / Brasil Pará; H. Andrade”, 1 specimen (MPEG); “Brasil Pará; Serra Norte; N2; Isca fruta; 22 a 24.I.1986 / Brasil Pará; M. F. Torres”, 1 specimen (MPEG); “Brasil Pará; Paragominas; Faz. Cachoeira; do Rio Vermelho; 18 a 21.I.1991 / Brasil Pará; P. Tadeu / Armadilha Malayse”, 1 specimen (MPEG); “Brasil Pará; Paragominas; Faz. Cachoeira; do Rio Vermelho; 18.I.1991 / Brasil Pará; P. Tadeu”, 1 specimen (MPEG); “Brasil Pará; Benevides; F. Morelândia; 6.III.1987 / Brasil Pará; J. Dias”, 1 specimen (MPEG); “Coleção Campos Seabra / Guajará; Borba Amazonas; Brasil IV.1943; A. Parko”, 1 specimen (MNRJ); “Brasil: AM; Itacoatiara; V/1962 Dirings”, 1 specimen (MZSP); “Forte Príncipe da; Beira. Rondônia; 19.XI-3.XII.1967; G. R. Kloss col. / Omalodes sp.; N. Degallier”, 1 specimen (MZSP).Diagnosis: Frons flat or slightly concave  and Ichilo (Bolivia). Mostly of the material was collected close to large rivers of those regions, especially in Pará, along the Caruaca river.Omalodes  gagatinus Erichson, 1847 and Omalodes  anthracinus Marseul, 1854, but this stria is a little longer on Omalodes  rivus. The punctures of the propygidium is reminiscent of that in Omalodes  lucidus Erichson, 1834, being completely covered by punctures which are sparser and weaker on the anterior half. The ninth tergite is more or less similar to all other species of the subgenus, but in Omalodes  rivus it is wider in the posterior half, and has a very subtle emargination along the anterolateral margin, this emargination is not known for any other species in the subgenus.This species resembles a few others in the subgenus; the frons and the length of the outer subhumeral stria is quite similar to There are few specimens with specific ecological data associated. Some labels indicate that the bait was fruit (orange or pineapple), probably rotten, since some other species of the genera are known to be attracted only by rotting vegetal or fruit material.http://zoobank.org/82F13716-A5D8-4541-AE55-CFA548C49ABFhttp://species-id.net/wiki/Omalodes_punctulatusHolotype: Male. BRAZIL: “15.IX.1955; Barueri; S. Paulo; 5876 / K. Lenko Leg. (MNRJ) Paratypes: BRAZIL: “Itapiranga; II 55”, 2 specimens (MAPA); PageBreak“Itapiranga; II 55”, 2 specimens (DZUP); “Est. Minas; Araguary; 5.XII.1921; Melim”, 1 specimen (MNRJ); “1225; 9.IX.1954; Barueri; Leg. K. Lenko”, 1 specimen (MNRJ); “29.X.1955; Barueri; S. Paulo; 6658 / K. Lenko Leg.”, 2 specimens (MNRJ); “Serro Azul; 3.40”, 1 specimen (MAPA); “1-44; Paraná; Monjolinho / 1628 / Coleção; F. Justus Jor.”, 1 specimen (DZUP); “Pelotas; II 55”, 1 specimen (DZUP).Frons punctate with a medial fovea ; dorsal PageBreakPageBreakPageBreakSize range: Length: 7–8mm, Width: 5–6mm. Body form  rivus, but spermatheca and spermathecal gland were not prepared for examination in this species.ody form : Oval, crm  punctulatus has a few punctures near the posterior margin of the elytra, and because of this, appears closely related to Omalodes  lucidus. However, its punctures are less numerous and more regular , while the punctures of Omalodes  lucidus are more numerous and irregularly distributed over the posterior margin of the elytra. Likewise the dorsal striae are weakly indicated, as in Omalodes  gagatinus and Omalodes  anthracinus, but when compared to those species, its striae are not so weakly indicated nor highly interrupted or shortened. The punctures of the propygidium resembles that of Omalodes  planifrons but it differs from this species by the punctures in the pronotum and its frons with a medial fovea. When compared to Omalodes  rivus this species has a shorter outer subhumeral stria, present only at posterior third and the frons has a medial fovea while in Omalodes  rivus the outer subhumeral stria is present in the posterior half and the frons is plan or only slightly concave.The inner subhumeral stria can be present only at the posterior fifth or absent; the punctures next to the posterior margin of the elytra can vary in number and intensity; the sutural stria can be completely absent and a few specimens have the dorsal striae more strongly indicated, although never as strongly as some other species of the genus.


In the 3,3′-diethyl-1,1′-di(1H-imidazol-3-ium) cation, the dihedral angle between the imidazole rings is 49.3 (1)°. In the [Ag2I4]2− anion, each AgI atom is bonded to three iodide anions, the two AgI atoms and two of the iodides forming Ag2I2 square-planar (r.m.s. deviation = 0.01 Å) units·The remaining two iodides, which are placed on opposite sides of the square, together with their centrosymmetric counterparts, link the square-planar Ag2I2 units into {[Ag2I4]2−}n polymeric chains via Ag—I bonds.The title compound, {(C DOI: 10.1107/S1600536812020715/lr2057Isup2.hklStructure factors: contains datablock(s) I. DOI:  crystallographic information; 3D view; checkCIF reportAdditional supplementary materials:


The most active compound towards urokinase was PhCH2SO2-d-Ser-Gly-Arg-OH with Ki value 5.4 μM and the most active compound toward thrombin was PhCH2SO2-d-Ser-NVa-Arg-OH with Ki value 0.82 μM. The peptides were nontoxic against porcine erythrocytes in vitro. PhCH2SO2-d-Ser-Gly-Arg-OH showed cytotoxic effect against DLD cell lines with IC50 values of 5 μM. For the highly selective determination of the interaction of some of the synthesised acids of tripeptides with urokinase and plasmin the Surface Plasmon Resonance Imaging sensor has been applied. These compounds bind to urokinase and plasmin in 0.05 mM concentration.Tripeptides of the general X-SO The proteolytic degradation of the extracellular matrix is essential for the processes of tissue remodelling. These processes take place in a number of distinct physiological events in the healthy organism such as matrix degradation, cell motility, angiogenesis and wound healing, as well as in the critical mechanisms in tumor invasion and metastasis. The plasminogen activation system has an important role as a proteolytic cascade in these degradation reactions  of urokinase. These peptides contains from 10 to 32 amino acids from this region of uPA  core -Arg-OH(NH2) n-NH2, n = 2–9, (CH2)m–OH, m = 2–4. H-d-Ser-Ala-Arg-NH-(CH2)5-NH2 inhibited urokinase with a Ki value of 6.3 μM. The third series of compounds with D-Ser-Ala-Arg sequence were N-sulfonylamides peptides -d-Ser-NVa-Arg-OH with a Ki value 0.85 μM 5-NH2) formula, where X = α-tolyl, p-tolyl, 2,4,6-triisopropylphenyl and AA = Ala, Gly, NVa , methanesulfonyl chloride, α-toluenesulfonyl chloride and 2,4,6-triisopropylbenzenesulfonyl chloride were purchased from Fluka . Fmoc-d-Ser(t-Bu)-OH (t-Bu = t-butyl) and 2-chlorotrityl chloride resin were purchased from Merck . TFA = trifluoroacetic acid, DIPEA = diisopropylethylamine, DIC = diisopropylcarbodiimide, piperidine, TBTU = tetrafluoroborate salt of the O-(benzotriazol-1-yl)-N,N,N′,N′-tetramethyluronium tetrafluoroborate, NMP = 1-methyl-2-pyrrolidone and 1,5-diaminopentanetrityl resin were obtained from Iris Biotech GmbH . DCM = dichloromethane, DMF = dimethyl-formamide and methanol were the products of Chempur . DCM was used without further purification. DMF was distillated over ninhydrin and stored under molecular sieves 4A. HPLC solvent acetonitrile was purchased from Merck . Urokinase, trypsin, kallikrein and Bzl-l-Arg-pNA.HCl (Bzl = benzyl) were purchased from Sigma . Plasmin, S-2444 (pyro-Glu-Gly-Arg-pNA.HCl), S-2238 (H-d-Phe-Pip-Arg-pNA), S-2251 , S-2266  were obtained from Chromogenix . Ac-Leu-Leu-Arg-H, cysteamine hydrochloride, N-(2-hydroxyethyl) piperazine-N′-(2-ethanesulfuric acid) (HEPES), N-ethyl-N′-(3-dimethylaminopropyl) carbodiimide (EDC) were purchased from Sigma . N-Hydroxysuccinimide (NHS) was obtained from Aldrich . Thrombin and phosphate buffered saline (PBS) were purchased from Lubelska Wytwórnia Szczepionek . t-PA was obtained from Boehringer Ingelheim GmbH . HBS-ES solution pH 7.4 , acetic buffer pH 3.79–5.57, phosphate buffer pH 7.17–8.04, carbonate buffer pH 8.50–9.86 , photopolimer ELPEMER SD 2054, hydrophobic protective paint SD 2368 UV SG-DG  were used as received. Aqueous solutions were prepared with MilliQ water (Simplicity® MILLIPORE).Fmoc-Arg(Pbf)-OH , chloranil, acetaldehyde, HOBt = 1-hydroxybenzotriazole, Fmoc-d-Ser(t-Bu)-OH molar ratio of amino acid/TBTU/HOBt/DIPEA/resin was 2:2:2:4:1. The sulfonyl chlorides were used as 5 M excess to resin and were dissolved in DCM with 10 M excess of DIPEA. The reactions were monitored with the Steward chloranil test  was used for analytical and semipreparatory HPLC . The major peak fraction was pooled and lyophilized. The molecular weight determination was performed by mass spectrometry using a Bruker Daltonics Esquire 6000  with electrospray ionization (ESI), (Table tris buffer—0.6 ml (pH 8.8),Determination of amidolytic activity was performed as previously described ,(b)tris buffer—0.5 ml (pH 8.4),synthetic substrate: S-2444 ;enzyme: thrombin (1 U/ml),(c)tris buffer—0.5 ml (pH 7.4),synthetic substrate: S-2238 ;enzyme: plasmin (0.4 U/ml),(d)borane buffer—0.5 ml (pH 7.5),synthetic substrate: S-2251 ;enzyme: trypsin (0.4 U/ml),l-Arg-pNA.HCl ;(e)tris buffer—0.6 ml (pH 9.0),synthetic substrate: Bzl-enzyme: kallikrein (3 U/ml),(f)tris buffer—0.6 ml (pH 8.4),synthetic substrate: S-2266 ;enzyme: t-PA (1.67 mg/ml),synthetic substrate: S-2288 .1–17) (as control 0.15 M NaCl). The mixture was incubated for 3 min at 37 °C, then the synthetic substrate was added. After 20 min of incubation, the reaction was stopped by adding 0.1 ml of 50 % acetic acid, and the absorbance of the released p-nitroaniline was measured at 405 nm . Every value represents the average of the triplicate determination. IC50 value was considered as the concentration of the inhibitor, which decreased the absorbance at 405 nm by 50 %, compared with the absorbance measured under the same conditions without the inhibitor. Ki was calculated from IC50 based on Cheng–Prusoff equation . The cells were maintained in DMEM supplemented with 5 % fetal bovine serum (FBS), 2 mmol/ml glutamine, 50 U/ml penicillin, 50 mg/ml streptomycin at 37 °C in a 5 % CO50 value was estimated from logarithm curves as shown in Table The toxicity of the evaluated peptides was determined by the method of Plumb et al.  in 10, 1g for 10 min to remove plasma and the buffy coat. The various concentrations of peptides  were incubated with the erythrocyte suspension for 1 h at 37 °C . After the centrifugation , 100 μl of the supernatant was transferred into sterilized 96-well plates, where hemoglobin release was monitored with the use of the Infinite M200 plate reader  by measuring the absorbance at 414 nm. Zero hemolysis (blank), hemolysis with Ac-Leu-Leu-Arg-H as reference compound for synthesised peptides and 100 % hemolysis which consisted of p-RBC suspended in PBS and 0.1 % Triton-X-100 were determined respectively. The percentage of hemolysis was calculated with the following formula: hemolysis (%) = (Abs414 nm in the peptide solution in PBS/Abs414nm in 0.1 % Triton-X-100 in PBS) × 100.Pig’s fresh red blood cells (p-RBC) were washed three times with PBS  and were centrifugated at 1,000Gold chips were manufactured as described in previous papers  and EDC (200 mM) was placed on the cysteamine-modified surface, and incubated at 37 °C for 1 h . In 2010 we synthesised N-sulfonylamide peptides with alanine as P2 5-NH2 were active urokinase inhibitors with Ki value 44.6 μM.We expected that the use of the specific tripeptide sequence towards urokinase (D-Ser-AA-Arg) would cause some selectivity 5-NH2 was the most active plasmin inhibitor with Ki value of 2 μM in this series of peptides. Compound 1 contains a fragment of pentamethylenediamine (cadaverine) as amide arginine residue in its structure which seems to have significant influence on the activity and selectivity. The moiety of cadaverine is the form of decarboxylated derivative of lysine. In literature, there are peptides with C-terminus lysine amide and also amide residue of lysine substituted with cadaverine, as inhibitors of plasmin  sensor.The most active compound towards urokinase was PhCH


Considerations have been given to compounds with activities ranging from “very active” to “weakly active”, leading to >500 chemical structures, mainly alkaloids, terpenoids, flavonoids, coumarins, phenolics, polyacetylenes, xanthones, quinones, steroids and lignans. However, only the compounds that showed anti-malarial activity, from “very active” to “moderately active”, are discussed in this review.Malaria is currently a public health concern in many countries in the world due to various factors which are not yet under check. Drug discovery projects targeting malaria often resort to natural sources in the search for lead compounds. A survey of the literature has led to a summary of the major findings regarding plant-derived compounds from African flora, which have shown anti-malarial/antiplasmodial activities, tested by  Plasmodium  furan-4,9-dione (70) and isopinnatal (71) from Kigelia pinnata (Bignoniaceae) . Isopinnatal (71) and the co-occurring kigelinol and isokigelinol exhibited lower activity against both strains. Bringmann et al. also reported that knipholone (68) and three of its natural derivatives from the same plant, as well as seven structurally related but simplified compounds, have been examined for their antiplasmodial activity against asexual erythrocytic stages of two strains of P. falciparum in vitro (K1/chloroquine-resistant and NF 54/chloroquine-sensitive) [Makinde  in mice . The bloelaceae) ,84, as wniaceae) . These cet al. [et al. [Vernonia brachycalyx (Asteraceae) and Toddalia asiatica (Rutaceae), respectively. Cubukcu et al. identified two isomeric 5-methylcoumarins from the roots of V. brachycalyx; 20-epi-cycloisobrachycoumarinone epoxide (72) and cycloisobrachycoumarinone epoxide (73), Table P. falciparum, showed that compound 72 was weakly active, with IC50 values of 160 μM and 54 μM, while for compound 73, the IC50 values were 111 μM and 54 μM, respectively. Noster et al. also isolated compound 73 from the ether extract of Exostema caribaeum but however only moderate activity [et al. isolated a new anti-malarial coumarin, 5,7-dimethoxy-8-(30-hydroxy-30-methyl- 10-butene) coumarin (74), from the roots of Toddalia asiatica[P. falciparum strains, with IC50 values of 16.2 μg mL−1 and 8.8 μg mL−1, respectively.Anti-malarial coumarins have been identified by Cubukcu et al.  and by N [et al.  from Verepoxide 7, Table 3activity . In addi asiatica. This co75) was isolated from the dichloromethane leaf extract of Schefflera umbellifera , harvested from Limpopo, South Africa by Mthembu et al. [in vitro against both the chloroquine-susceptible (D10) and chloroquine-resistant (K-1) strains of P. falciparum for anti-malarial activity, with an IC50 value of 28.2 μg mL−1.The anti-malarial coumarin 7-hydroxy6-methoxycoumarin or scopoletin  and 3-hydroxy-5-methoxyxanthone (77) were isolated from stem bark of Hypericum lanceolatum (Hypericaceae) from Cameroon by Zofou et al. [50 values of 13.56 μg mL−1, and 8.28 μg mL−1, respectively, on the multidrug-resistant W2mef strain of P. falciparum. Six other anti-malarial xanthones were isolated from the methanol extract of the stem bark of Allanblackia monticola (Guttiferae) from Cameroon, by Azebaze et al. [78), garciniafuran (79), tovophyllin A (80), rubraxanthone (81), norcowanin (82) and mangostin (83), Figure 50 of 1.3 μM on FcM29 and an IC50 of 6.9 μM on F32. The molecules with interesting activities are known to be norcowanin (IC50 of 6.3 μM on F32) and mangostin , symphonin (85) and globuliferin (86) [Pentadesma butyracea, using the antiplasmodial test, led to the isolation of a new bioactive xanthone, named pentadexanthone (87) (IC50 = 3 μM against W2), together with three known compounds: cratoxylone (88) (IC50 = 2.89 μM), α-mangostin (89) (IC50 = 2.77 μM), and garcinone E (90) (IC50 = 0.41 μM) [The anti-malarial xanthones; 5-hydroxy-3- methoxyxanthone  . Bioassa0.41 μM) .91), isolated from the aerial parts of Ajuga remota, exhibited high antiplasmodial activity against the chloroquine-sensitive FCA 20/GHA strain of P. falciparum, with an IC50 value of 8.2 μM [Vernonia amygdalina[et al. reported the isolation of vernonioside A1 (92), A2 (93), A3 (94), A4 (95) and B1 (96), Figure P. falciparum, with IC50 values of 139.7, 94.1, 245.1, 81.8 and 46.1 μg mL−1, respectively [V. amygdalina.The steroid, ergosterol-5,8-endoperoxide , A3 9, A4 95)91), isolPycnanthus angolensis (Myristicaceae) is a plant used in traditional medicine against several diseases. Its bark has been used to treat fever and malaria in São Tomé and Príncipe islands. Ramalhete et al. submitted the dichloromethane extract of the bark to anti-malarial screening and observed an activity against 3D7 P. falciparum strain (IC50 = 1.6 μg mL−1) [97), (−)-dihydroguaiaretic acid (98), 4′-hydroxy-3,3′,4-trimethoxylignan (99), 4,4′-diacetyl-3,3′-dimethoxylignan (100), talaumidin (101), hinokinin (102), and heliobuphthalmin (103), Figure in vitro anti-malarial activity of the compounds was evaluated against 3D7 and Dd2 P. falciparum strains. The best in vitro activities were exhibited by compound 97 against the 3D7 strain (IC50 = 31.0 μg mL−1) and by compound 101 against the Dd2 strain (IC50 = 20.7 μg mL−1).μg mL−1) . This waxylignan 0, talaumAsparagus africanus (Asparagaceae) is used by the Akamba tribe in Kenya to treat malaria. A bioassay-guided fractionation of the root extract led to the isolation of the lignan nyasol (104), along with the sapogenin muzanzagenin (119), Figure P. falciparum schizonts with the IC50 of 49 μM, while muzanzagenin showed a moderate in vitro activity against four different malaria schizont strains the IC50 values were 16, 163, 23, and 16 μM, respectively.nzagenin 9, Figure105), derived from the ethyl acetate extract of aerial parts of Lippia javanicais, harvested from South Africa, was shown to be active against the chloroquine-sensitive D10 strain of P. falciparum with an IC50 value of 9.1 μg mL−1, and is also mildly cytotoxic [106) was derived from extracts of the whole plant of Helichrysum cymosum (Asteraceae) from South Africa by Jakupovic et al.[et al.[Lippialactone  showed in vitro activity (IC50 ~ 3 μg mL-1) against the chloroquine-sensitive D10 and the chloroquine-resistant (K-1) strains of P. falciparum[et al. further investigated the extract by bioassay-guided fractionation and two structurally related hirsutinolides displaying in vitro antiplasmodial activity (IC50 ~ 0.2 μg mL−1 against D10) were isolated. These were 8α-(2- methylacryloyloxy)-3-oxo-1-desoxy-1,2-dehydrohirsutinolide-13-O-acetate (107), and 8α- (5′-acetoxysenecioyloxy)-3-oxo-1-desoxy-1,2-dehydrohirsutinolide-13-O-acetate (108). These were found to be cytotoxic to mammalian Chinese hamster ovarian (CHO) cells at similar concentrations, but proved to be attractive scaffolds for structure-activity relationship studies. Two main privileged substructures, a 2(5H)-furanone unit and a dihydrofuran-4- one unit, were identified as potential pharmacophores, which may be responsible for the observed biological activity. Mucochloric and mucobromic acids were selected as appropriate 2(5H)-furanone substructures and these were shown to have comparable activity against the D10 and superior activity against the K1 strains relative to the hirsutinolide natural product. Mucochloric and mucobromic acids (109 and 110) also showed selective cytotoxicity to the malaria parasites compared to mammalian (CHO) cells in vitro. The antiplasmodial data obtained with respect to these two acids suggest that the 2(5H)- furanone substructure is a key pharmacophore in the observed antiplasmodial activity. The identification of antiplasmodial hirsutinolides from V. staehelinoides suggests that they may play a role in the medicinal properties of the plant, but their potential for the development of anti-malarial drugs is limited due to inherent cytotoxicity and lack of selectivity. The results did however lead to the identification of potential pharmacophores, a 2(5H)-furanone unit and a dihydrofuran-4-one unit.The dichloromethane extract of the leaves of alciparum. Pillay 111), from the stem bark of Hypericum lanceolatum (Hypericaceae), exhibited an interesting activity against the multidrug-resistant strain W2mef, with an IC50 of 13.56 μg mL−1[et al. isolated guttiferone A (112) from the seeds of Symphonia globulifera (Clusiaceae) [Plasmodium sp. with IC50 of 3.17 μM. Mulberrofuran F (113), from the stem bark of Morus mesozygia (Moraceae), was active against FcB1-Columbia strain, considered to be resistant against chloroquine, with IC50 of 2.6 μg mL−1[114) and specicoside (115), derived from the stem bark extract of Kigelia africana (Bignoniaceae) [P. falciparum with respective IC50 values of 0.67 μg mL−1 and 0.52 μg mL−1.The benzophenone 2,5,2′,6′-tetrahydroxybenzophenone (6 μg mL−1. Ngouelasiaceae) . This co6 μg mL−1. Atranorniaceae) ,69, were116) and 2-(1′β-geranyl-5′β-hydroxy-2′- oxocyclohex-3′-enyl) acetic acid (117) were isolated by Mbah et al. from the leaves of Glossocalyx brevipes (Monimiaceae) [116 and 117 exhibited both anti-malarial [118) and the saponin spiroconazole A (119), respectively isolated from Asparagus africanus (Asparagaceae) [Dracaena sp. (Dracaenaceae) [The homogentisic acid derivatives methyl 2-(1′β-geranyl-5′β-hydroxy-2′- oxocyclohex-3′-enyl) acetate (miaceae) . Compounmalarial  and antimalarial . The sapagaceae)  and fromenaceae) , also deP. falciparum. With multiple resistance developed by the malaria parasite, the cry has been towards obtaining new effective drugs. Attempts to develop ‘green pharmacies’ for improved phytomedicines against malaria are being encouraged by some NGOs and governments as part of their efforts to control malaria [In this review, an attempt has been made to summarise the main finding of several research groups engaged in the search for naturally occurring active principles from African medicinal plants against  malaria . Additio malaria  and it h malaria . The aut malaria -110 and  malaria . Since t malaria -116, eff malaria ,118. TheADMET: Absorption, distribution, metabolism, excretion and toxicity; ATM: African traditional medicine; DMPK: Drug metabolism and pharmacokinetics; FAS II: Fatty acid sytem II; NP: Natural product; WHO: World Health Organization; WM: Western medicine.The authors declare that they have no competing interests.FNK, LLL, JCN, and LMM conceived the idea. FNK, LLL and PAO participated in data collection. FNK and PAO contributed to data analysis, discussion of results and the conception of the paper under the supervision of LMM, WS, LLL, and JCN. FNK and PAO wrote the first draft of the paper and all authors agreed on the final version before submission.


The anion is composed of a bidentate coordinated 3,4,5-trioxocyclopent-1-ene-1,2-dithiolate (dtcroc) group forming a distorted tetrahedral configuration around the CdII ion. The dihedral angle between the least-squares planes of the ten-atom sulfur-substituted croconate groups in the anion is 84.10 (8)°. The crystal packing is stabilized by weak C—H⋯O and C—H⋯S cation–anion hydrogen-bond interactions. In each of the two cations one butyl group is disordered over  two positions in the ratios 0.589 (11):0.411 (11) and 0.796 (12):0.204 (12).The title compound, (C DOI: 10.1107/S1600536810049913/jj2066Isup2.hkl            Structure factors: contains datablocks I. DOI:  crystallographic information; 3D view; checkCIF report            Additional supplementary materials:


The coordination geometry is defined by three N-atom donors from the tridentate 2-(pyridin-2-yl)-N-(pyridin-2-ylmethylidene)ethanamine ligand and two terminal Cl atoms. Although the H atoms of the lattice water molecule were not located, O⋯O distances of 3.103 (7) Å and O⋯Cl distances of 3.240 (3) and 3.482 (4) Å suggest that hydrogen bonding is responsible for the stabilization of the crystal packing.In the title complex, [MnCl DOI: 10.1107/S1600536812037877/fk2065Isup2.hklStructure factors: contains datablock(s) I. DOI:  crystallographic information; 3D view; checkCIF reportAdditional supplementary materials:


II ion in the title complex, [PdCl2(C14H10N4)], has a slightly distorted square-planar environment defined by the two pyridine N atoms of the chelating 2,3-di-2-pyridylpyrazine ligand and two chloride anions. The pyridine rings are considerably inclined to the least-squares plane of the PdCl2N2 unit [maximum deviation = 0.073 (1) Å], with dihedral angles of 64.19 (9) and 66.55 (9)°. The pyrazine ring is almost perpendicular to this plane and the dihedral angle is 88.2 (1)°. Two independent intermolecular C—H⋯Cl hydrogen bonds, both involving the same Cl atom as a hydrogen-bond acceptor, give rise to chains running along the a and b axes, generating a layer structure extending parallel to (001). Molecules are stacked in columns along the a axis. Along the b axis, successive molecules stack in opposite directions.The Pd DOI: 10.1107/S1600536811043753/ng5256Isup2.hkl            Structure factors: contains datablock(s) I. DOI:  crystallographic information; 3D view; checkCIF report            Additional supplementary materials:


A terminal Br atom and a bidentate chelating 4,7-diphenyl-1,10-phenanthroline ligand complete a square-pyramidal geometry for the CdII atom. In the crystal, C—H⋯Br hydrogen bonds and π–π contacts between the pyridine and phenyl rings [centroid–centroid distances = 3.704 (4) and 3.715 (4) Å] lead to a three-dimensional supramolecular structure.The title compound, [Cd DOI: Click here for additional data file.10.1107/S1600536812045928/hy2601Isup2.hklStructure factors: contains datablock(s) I. DOI:  crystallographic information; 3D view; checkCIF reportAdditional supplementary materials:


The orientation of the molecules in the orthorhombic cell shows that the structure can not be described in the space group Pnma, which has the same systematic absence conditions. The long inner-cluster C—C distance of 1.510 (5) Å is typical for {1,2-Me2-closo-1,2-C2B10} derivatives. The asymmetric unit of the title compound, C DOI: 10.1107/S1600536810022440/si2268Isup2.hkl            Structure factors: contains datablocks I. DOI:  crystallographic information; 3D view; checkCIF report            Additional supplementary materials:


This molecule looks like a space shuttle, the IDA ligand is its empennage (tail), and the 4-OXPy ligand is its airframe. The complexes are linked into two-dimensional supramolecular layers parallel to (100) by three pairs of O—H⋯O hydrogen bonds. Two pairs of N—H⋯O hydrogen bonds further connect these supramolecular layers, forming a three-dimensional supramolecular network. In the title complex, [Cu(C N-heterocyclic ligand, see: Roman-Alpiste et al. , IDA and an  al. 1999; Kundu e al. 2005; Chen et al. 1990; Zhang e al. 2008; Selvaku al. 2006; Siddiqi al. 2009; Setha e al. 2010; Campos  al. 1996; Castine al. 1995; Brandi- al. 2003; Craven  al. 2003. For hyd al. 2003. For theSpek 2009.       4H5NO4)(C6H6N2O)(H2O)] = 0.041                           wR(F                           2) = 0.095                           S = 1.04                           2739 reflections229 parameters3 restraintsH atoms treated by a mixture of independent and constrained refinementmax = 0.57 e Å−3                        Δρmin = −0.50 e Å−3                        Δρ                                 PROCESS-AUTO  I, global. DOI: 10.1107/S1600536811033459/zk2022Isup2.hkl            Structure factors: contains datablock(s) I. DOI:  crystallographic information; 3D view; checkCIF report            Additional supplementary materials:


The TiIV atom is in a pseudo-octahedral coordination environment with the imidazolido-group N-atoms occupying apical positions and amido- and imino-N-atoms cis-located in the equatorial plane. The presence of two bidentate chelating ligands determines the chirality of the TiIV atom. The crystallographically independent unit, except for its phenyl rings, adopts nearly pseudo-C               2 symmetry (rotation around a twofold axis passing through the Ti atom and the centre of the imino-N⋯imino-N segment). The Ti—Namido, Ti—Nimidazolido, and Ti—Nimino bond lengths essentially differ, increasing by approximately 0.2 Å in the series. All ligating N atoms are in a nearly planar environment, which is indicative of additional pπ–dπ donations towards the metal atom. The two diazametallacyclic units are planar within 0.03 and 0.05 Å.The title compound, [[Ti(C IV complexes bearing two chelating amido-imino and two amido ligands see: Xiang et al. 2(C11H10N3)2] = 0.044                           wR(F                           2) = 0.100                           S = 0.94                           5478 reflections356 parametersH-atom parameters constrainedmax = 0.27 e Å−3                        Δρmin = −0.25 e Å−3                        Δρ                                 APEX2  used to solve structure: SHELXS97 (Sheldrick, 2008SHELXL97 (Sheldrick, 2008SHELXTL (Sheldrick, 2008OLEX2 (Dolomanov et al., 2009SHELXTL and OLEX2.Data collection: 10.1107/S1600536811013183/dn2674sup1.cif            Crystal structure: contains datablocks I, global. DOI: 10.1107/S1600536811013183/dn2674Isup2.hkl            Structure factors: contains datablocks I. DOI:  crystallographic information; 3D view; checkCIF report            Additional supplementary materials:


III atom in the title complex, [CoCl2(C40H48N4)]ClO4, is octahedrally coordinated within a trans-Cl2N4 donor set provided by the tetradentate macrocylic ligand and two chloride ions. The N—H atoms, which are orientated to one side of the N4 plane, form hydrogen bonds with chloride ions and perchlorate-O atoms. These along with C—H⋯O interactions consolidate the three-dimensional crystal structure. One of the benzene rings was disordered. This was resolved over two positions with the major component of the disorder having a site-occupancy factor of 0.672 (4).The Co N-substituted derivatives and metal complexes, see: Bembi et al. ]ClO4                         = 0.048                           wR(F                           2) = 0.142                           S = 1.05                           7470 reflections476 parameters2 restraintsH atoms treated by a mixture of independent and constrained refinementmax = 0.72 e Å−3                        Δρmin = −0.50 e Å−3                        Δρ                                 CrystalClear  used to solve structure: PATTY in DIRDIF92  global, I. DOI: 10.1107/S1600536811046484/pv2479Isup2.hkl            Structure factors: contains datablock(s) I. DOI:  crystallographic information; 3D view; checkCIF report            Additional supplementary materials:


II               8Si6O1510 , is a phyllosilicate mineral and a member of the pyrosmalite series 8Si6O1510, which includes pyrosmalite-(Mn), as well as friedelite and mcgillite, two polytypes of pyrosmalite-(Mn). This study presents the first structure determination of pyrosmalite-(Fe) based on single-crystal X-ray diffraction data from a natural sample from Burguillos del Cerro, Badajos, Spain. Pyrosmalite-(Fe) is isotypic with pyrosmalite-(Mn) and its structure is characterized by a stacking of brucite-type layers of FeO6-octahedra alternating with sheets of SiO4 tetrahedra along [001]. These sheets consist of 12-, six- and four-membered rings of tetrahedra in a 1:2:3 ratio. In contrast to previous studies on pyrosmalite-(Mn), which all assumed that Cl and one of the four OH-groups occupy the same site, our data on pyrosmalite-(Fe) revealed a split-site structure model with Cl and OH occupying distinct sites. Furthermore, our study appears to suggest the presence of disordered structural water in pyrosmalite-(Fe), consistent with infrared spectroscopic data measured from the same sample. Weak hydrogen bonding between the ordered OH-groups that are part of the brucite-type layers and the terminal silicate O atoms is present.Pyrosmalite-(Fe), ideally Fe For oth al. 1993; Stillwe al. 1993; Takéuch al. 1963, 1969 ▶;8Si6O15(OH0.814Cl0.186)10·0.45H2OFeM                           r = 1067.35                           Trigonal, a = 13.3165 (2) Å                           c = 7.0845 (2) Å                           V = 1087.98 (4) Å3                                                   Z = 2                           Kα radiationMo −1                        μ = 5.85 mmT = 293 K                           0.09 × 0.08 × 0.08 mm                                 Bruker APEXII CCD area-detector diffractometerSADABS; Sheldrick, 2005T                           min = 0.622, T                           max = 0.653Absorption correction: multi-scan (13410 measured reflections1476 independent reflectionsI > 2σ(I)1141 reflections with R                           int = 0.032                                                            R[F                           2 > 2σ(F                           2)] = 0.023                           wR(F                           2) = 0.068                           S = 1.05                           1476 reflections90 parametersAll H-atom parameters refinedmax = 0.65 e Å−3                        Δρmin = −0.56 e Å−3                        Δρ                                 APEX2  used to solve structure: SHELXS97  I, global. DOI: 10.1107/S1600536811052822/wm2570Isup2.hkl            Structure factors: contains datablock(s) I. DOI:  crystallographic information; 3D view; checkCIF report            Additional supplementary materials:


R-module A is called GF-regular if, for each a ∈ A and r ∈ R, there exist t ∈ R and a positive integer n such that rntrna = rna. We proved that each unitary R-module A contains a unique maximal GF-regular submodule, which we denoted by MGF(A). Furthermore, the radical properties of A are investigated; we proved that if A is an R-module and K is a submodule of A, then MGF(K) = K∩MGF(A). Moreover, if A is projective, then MGF(A) is a G-pure submodule of A and MGF(A) = M(R) · A.An  R is a commutative ring with identity and all modules are left unitary, unless otherwise stated. Recall that an element r ∈ R is said to be regular if there exists t ∈ R such that rtr = r; a ring R is called regular if and only if each element of R is regular. An ideal I of a ring R is regular if each of its elements is regular in R; indeed, a regular ideal I of R is itself a regular ring [R contains a unique maximal regular ideal M(R) which satisfies the well-known radical properties. The ideal M(R) is called the regular radical of R.Throughout this paper, lar ring . Brown alar ring  that eacF-regular modules  was genAlso, in  the concGF-regular submodule, which we denote by MGF(A), and we show that MGF(A) satisfies some but not all of the usual radical properties.In this paper we show that each module contains a unique maximal R be any ring. Every R-module contains a unique maximal GF-regular submodule. Let R be any ring, let A be an R-module, and letG ≠ ϕ because (0) is a GF-regular submodule of A. Let {Ni} be an ascending chain in G and B = ⋃i∈ΛNi. Let b ∈ B; there exists j ∈ Λ such that b ∈ Nj, but Nj is a GF-regular submodule; then; for each r ∈ R, there exist t ∈ R and a positive integer n such that rntrnb = rnb; therefore b is a GF-regular element in B which implies that B is a GF-regular R-module. Now, by Zorn's lemma, G contains a maximal element which we call MGF. To prove the uniqueness of MGF, assume that MGF1 and MGF2 be two maximal GF-regular submodules in A; then for any maximal ideal P of R each of MGF1p and MGF2p is semisimple over Rp [MGF1p∩MGF2p = Kp; then Kp⊆MGF1p and Kp⊆MGF2p; thus MGF1p = Kp + A1p and MGF2p = Kp + A2p, where A1p and A2p are two submodules of Ap [MGF1p + MGF2p = A1p + Kp + A2p, but each of A1p, A2p, and Kp is a semisimple submodule; thus MGF1p + MGF2p is a semisimple submodule which implies that MGF1p + MGF2p is GF-regular [MGF1 + MGF2 is a GF-regular submodule [MGF1 and MGF2 is a maximal GF-regular submodule and hence MGF1 + MGF2 = MGF2 = MGF1.  Let s of Ap . Hence, GF-regular submodule of an R-module A by MGF(A). It is obvious that MGF(A) contains every GF-regular submodule of A; this means that MGF(A) is a GF-regular submodule which is not contained properly in any other GF-regular submodule. In fact, MGF(A) is the sum of all GF-regular submodules of A and MGF(A) = A if and only if A is a GF-regular module. We denote the unique maximal Z-module Zn is GF-regular for each positive integer n [MGF(Zn) = Zn. (a) Since the nteger n , then MZ-module Q is not GF-regular [MGF(Q) = (0). (b) Each element in the -regular ; hence Mp be a prime number and let A = Zp∞ = ⋃i∀Zpi be a Z-module. Let a ∈ ⋃i∀Zpi; then there exists a positive integer m such that a ∈ Zpm, but Zn is a GF-regular Z-module for each positive integer n; hence a is a GF-regular element, so a ∈ MGF(Zp∞) which implies that MGF(Zp∞) = Zp∞. (c) Let Q/Z be a Z-module; since Q/Z is a torsion Z-module, then Q/Z is a GF-regular Z-module [Q/Z = ∑pZp∞ for each prime number p, then MGF(Q/Z) = ∑pZp∞ for all primes p. (d) Let A and B be R-modules, and let K be a submodule of A; then MGF(K) = K∩MGF(A), MGF(A ⊕ B)⊆MGF(A) ⊕ MGF(B). Let K be a submodule of A, and let k ∈ MGF(K); then k ∈ K and k is GF-regular in K which implies that k is GF-regular in A, thus k ∈ K∩MGF(A). Conversely, let k ∈ K and k ∈ MGF(A); therefore k is GF-regular in K which means that k ∈ MGF(K) and hence MGF(K) = K∩MGF(A). (a) Let c ∈ MGF(A ⊕ B); then c = , where a ∈ A and b ∈ B. Since c is GF-regular, then each of a and b is GF-regular which means that a ∈ MGF(A) and b ∈ MGF(B); hence c ∈ MGF(A) ⊕ MGF(B). (b) Let A and A′ be R-modules, and let f : A → A′ be an R-homomorphism; then f(MGF(A))⊆MGF(f(A)). Let a ∈ MGF(A), then R/ann(a) is a π-regular ring, but ann(a)⊆ann(f(a)); thus R/ann(f(a)) is an epimorphic image of R/ann(A); hence it is a π-regular ring. Therefore, f(a) ∈ MGF(f(a)) and f(MGF(A))⊆MGF(f(A)).  If f : A → A′ is an R-epimorphism, then f(MGF(A)) ≠ MGF(A′) in general. In fact, let π : Z → Z4≃Z/4Z be the natural map, where Z and Z4 are Z-modules. It is easy to check that MGF(Z) = (0), f(MGF(Z)) = (0), but MGF(Z4)≃Z2. (a) If R, M(R/M(R)) = (0) which is not true in case of GF-regular modules; this means that MGF(A/MGF(A))≠(0) (as in (a)). (b) It is shown in  that, foR-module A, M(R) · A⊆MGF(A). For each a ∈ A, let f : R → A be an R-homomorphism defined by f(r) = ra. Then f(M(R))⊆MGF(A) by M(R) · A = ∑af(M(R)); hence M(R) · A⊆MGF(A). For each J(R) be the Jacobson radical of a ring R. Brown and McCoy proved in [M(R)∩J(R) = (0). However, this is not true for GF-regular modules; for example, if A = Z4 is a Z-module, then MGF(A)≃Z2, J(A)≃Z2 and MGF(A)∩J(A)≠(0).Let roved in  that M; if P is a finitely generated G-pure submodule of an R-module A such that P⊆RrnA for some positive integer n, then P = 0. Let P = RrnP and by Nakayama's lemma [P = 0.  By 's lemma , P = 0. A be an R-module. If MGF(A) is a G-pure submodule of A, then MGF(A)∩J(R) · A = (0). Let r ∈ MGF(A)∩J(R) · A, and let P = Rr. It is clear that P⊆MGF(A). Since MGF(A) is a GF-regular module, then P is a G-pure submodule in MGF(A) [MGF(A) is G-pure in A; hence P is G-pure in A. Now, P⊆J(R) · A, so P = 0 by MGF(A)∩J(R) · A = (0). Let M(R) is always a pure ideal in R. Hence M(R) is G-pure [Recall that s G-pure .A be a projective R-module; then MGF(A) = M(R) · A, MGF(A) is a G-pure submodule of A, MGF(A)∩J(A) = (0). Let a ∈ A we have that a = ∑ifi(a)ai, where a, ai ∈ A for all i and fi ∈ A* : = HomR. If a ∈ MGF(A), then the submodule Ra is GF-regular and fi(Ra) is a GF-regular ideal in R by M(R). Thus MGF(A)⊆M(R) · A. We get the other direction of the inclusion by (a) By the dual basis lemma , for eacK and L of R, (K ∩ L)A = KA ∩ LA; it is enough to show this locally; thus we may assume that A is free. It is clear that (K∩L)A⊆KA∩LA. On the other hand, let x ∈ KA∩LA; thenri = si and x ∈ (K∩L)A. Now, let K be any ideal in R; then by (a) we get MGF(A) ∩ KA = M(R) ∩ KA = (M(R) ∩ K)A. But M(R) is G-pure ideal, so, for each r ∈ R, there exists a positive integer n such that M(R)∩Rrn = RrnM(R); hence MGF(A)∩RrnA = RrnM(R) · A = RrnMGF(A). (b) First we claim that; for any two ideals A is projective, then J(A) = J(R) · A [MGF(A)∩  J(A) = (0) by (c) Since J(R) · A  which imR be any ring, and let A be any R-module such that MGF(A) is a G-pure submodule and J(A) = J(R) · A; then MGF(A)∩J(A) = (0). Let J(A) = J(R) · A, then by MGF(A)∩J(A) = (0). Since A is a GF-regular R-module, then MGF(A) = A; hence J(R) · A = J(R) · A∩A = J(R) · A∩MGF(A) = (0). In fact, this shows that If π-regular ring is GF-regular, but the converse need not be true in general. The next result shows how the converse may be true, but first we recall that if A is an R-module, then the trace of A is tr(A) = ∑f∈A*f(A), where A* = Hom.In  we noticA be a GF-regular R-module. If tr(A) = R, then R is a π-regular ring. Let a ∈ A and f ∈ A* = Hom, since Ra is a GF-regular submodule of A, then by [f(Ra) is a π-regular ideal. Thus f(Ra)⊆M(R), but f(Ra)⊆tr(A); hence tr(A) = R⊆M(R), which implies that M(R) = R and R is π-regular.For each


The ZnII ion is coordinated in a distorted tetrahedral environment by two N atoms from the chelating 2,9-dimethyl-1,10-phenanthroline ligand and two bromide ions. There is intermolecular π–π stacking between adjacent phenanthroline units, with centroid–centroid distances of 3.594 (3) and 3.652 (3) Å.The reaction of equimolar amounts of zinc bromide and 2,9-dimethyl-1,10-phenanthroline in dry methanol provided the title compound, [ZnBr DOI: 10.1107/S1600536812022738/bt5921Isup2.hklStructure factors: contains datablock(s) I. DOI:  crystallographic information; 3D view; checkCIF reportAdditional supplementary materials:


The ZnII atom is coordinated by three O and one N atom from one 2-(2H-benzotriazol-2-yl)-4,6-di-tert-pentylphenolate ligand and two bridging trimethylsilanolate anions in a distorted tetrahedral geometry. The dihedral angle between the benzotriazole ring system and the benzene ring is 19.83 (5)°. The tert-pentyl groups are disordered over two orientations with refined site-occupancy ratios of 0.858 (4):0.142 (4) and 0.665 (6):0.335 (6).The binuclear title complex, [Zn DOI: 10.1107/S1600536811054134/rz2685Isup2.hkl            Structure factors: contains datablock(s) I. DOI:  crystallographic information; 3D view; checkCIF report            Additional supplementary materials:


Matrix metalloproteinase-9 (MMP-9) has been shown to be induced by cytokines including TNF-α and may contribute to bone inflammatory diseases. However, the mechanisms underlying MMP-9 expression induced by TNF-α in MC3T3-E1 cells remain unclear.We applied gelatin zymography, Western blot, RT-PCR, real-time PCR, selective pharmacological inhibitors of transcription , translation , c-Src (PP1), MEK1/2 (U0126), p38 MAPK (SB202190), JNK1/2 (SP600125), and NF-κB (Bay11-7082), respective siRNAs transfection, promoter assay, immunofluorescence staining, and ELISA to investigate the MMP-9 expression and soluble ICAM-1 (sICAM-1) release induced by TNF-α in MC3T3-E1 cells. Here we demonstrated that TNF-α-induced MMP-9 expression was attenuated by Act.D, CHI, PP1, U0126, SB202190, SP600125, and Bay11-7082, and by the transfection with siRNAs for ERK2, p38 MAPK, and JNK2. TNF-α-stimulated TNFR1, TRAF2, and c-Src complex formation was revealed by immunoprecipitation and Western blot. Furthermore, TNF-α-stimulated NF-κB phosphorylation and translocation were blocked by Bay11-7082, but not by PP1, U0126, SB202190, or SP600125. TNF-α time-dependently induced MMP-9 promoter activity which was also inhibited by PP1, U0126, SB202190, SP600125, or Bay11-7082. Up-regulation of MMP-9 was associated with the release of sICAM-1 into the cultured medium, which was attenuated by the pretreatment with MMP-2/9i, an MMP-9 inhibitor.In this study, we demonstrated that TNF-α up-regulates MMP-9 expression via c-Src, MAPKs, and NF-κB pathways. In addition, TNF-α-induced MMP-9 expression may contribute to the production of sICAM-1 by MC3T3-E1 cells. The interplay between MMP-9 expression and sICAM-1 release may exert an important role in the regulation of bone inflammatory diseases. Matrix metalloproteinases (MMPs) are a family of extracellular matrix (ECM)-degrading enzymes and induced by different stimuli including growth factors, cytokines, and tumor promoters . MMPs plSeveral proinflammatory mediators, including tumor necrosis factor-α (TNF-α) have been reported to be associated with many bone functions such as resorption and inflammation. The expression of MMPs has been shown to be regulated by several extracellular stimuli such as TNF-α and IL-1β in various cell types ,10. Numein vitro. In. In25]. in vitro. TNF-α sin vitro. The levin vitro,31. Howei.e. ERK1/2, p38 MAPK, and JNK1/2) and NF-κB is essential for the induction of the MMP-9 gene expression in these cells. Moreover, the induction and activation of MMP-9 are important for sICAM-1 release from MC3T3-E1 cells. These results provide new insights into the mechanisms of TNF-α action that the c-Src-dependent MAPKs and IKK/NF-κB may be associated with the MMP-9 up-regulation and the sICAM-1 release from osteoblasts-like MC3T3-E1 cells.In this study, the mechanisms underlying TNF-α-induced MMP-9 expression and the effects of increased MMP-9 on MC3T3-E1 cells were investigated. We found that the activation of three MAPKs (536), anti-phospho-c-Src, anti-phospho-ERK1/2, anti-phospho-p38 MAPK, anti-phospho-JNK1/2, and anti-phospho-IκB-α antibodies were from Cell Signaling . anti-NF-κB (p65), anti-lamin A, anti-TRAF2, anti-TNFR1, anti-c-Src, anti-ERK2, anti-p38, anti-JNK2, anti-IκB-α, and anti-sICAM-1 antibodies were from Santa Cruz . The anti-GAPDH antibody was from Biogenesis . Actinomycin D (Act.D), cycloheximide (CHI), PP1, U0126, SB202190, SP600125, GM6001, MMP2/9 inhibitor, and Bay11-7082 were from Biomol . Enzymes and other chemicals were from Sigma .Minimal essential medium-alpha (α-MEM), fetal bovine serum (FBS), and TRIzol were purchased from Invitrogen . Hybond C membrane and ECL Western blotting detection system were from Amersham Biosciences . Recombinant human TNF-α and the anti-TNFR1 neutralizing antibody were from R&D System . Luciferase assay kit was from Promega . Metafectene transfection reagent was from Biontex Lab . SMARTpool RNA duplexes corresponding to Src, TRAF2, ERK2, p38 MAPK, JNK2, and scrambled #2 siRNA were from Dharmacon Research Inc . Anti-phospho-IKKα/β, anti-phospho-NF-κB p65 (Ser2 atmosphere. When the cultures reach confluence (4 days), cells were treated with 0.05% (w/v) trypsin/0.53 mM EDTA for 5 min at 37°C. The cell suspension was diluted with α-MEM containing 10% (v/v) FBS to a concentration of 2 × 105 cells/ml. The cell suspension was plated onto (1 ml/well) 12-well culture plates and (10 ml/dish) 10-cm culture dishes for the measurement of protein expression and mRNA accumulation, respectively. Culture medium was changed after 24 h and then every 3 days.Murine osteoblastic cell line, MC3T3-E1, a homogeneous source of non-transformed cell, was a gift from Dr. Hyun-Mo Ryoo  . MC3T3-EMC3T3-E1 cells were plated onto 12-well culture plates and made quiescent at confluence by incubation in serum-free α-MEM for 24 h. Growth-arrested cells were incubated with TNF-α at 37°C for the indicated time intervals. When inhibitors were used, they were added 1 h prior to the application of TNF-α. The culture medium was collected and centrifuged at 14000 rpm for 5 min at 4°C to remove cells and debris, then each sample was mixed with equal amounts of (×2) non-reducible sample buffer and electrophoresed on 10% SDS-PAGE containing 1 mg/ml gelatin as the protease substrate, as previously described .g at 4°C for 1 h to yield the whole cell extract [Growth-arrested MC3T3-E1 cells were incubated with TNF-α at 37°C for the indicated time intervals. The cells were washed, scraped, collected, lysed with ice-cold lysis buffer, and centrifuged at 45,000  extract . SamplesTotal RNA was isolated from MC3T3-E1 cells treated with TNF-α for the indicated time intervals with TRIzol according to the protocol of the manufacturer. RNA concentration was spectrophotometrically determined at 260 nm. First-strand cDNA synthesis was performed with 2 μg of total RNA using random hexamers as primers in a final volume of 20 μl . The reaction was carried out at 37°C for 60 min. cDNAs encoding β-actin and MMP-9 were amplified from 3 to 5 μl of the cDNA reaction mixture using specific gene primers. Oligonucleotide primers for β-actin and MMP-9 were follow as: β-actin (514 bp): 5′-GAACCCTAAGGCCAACCGTG-3′ (sense), 5′-TGGCATAGAGGTCTTTACGG-3′ (antisense); MMP-9 (458 bp): 5′-TGTTCAGCAAGGGGCGTGTC-3′ (sense) 5′-AAACAGTCCAACAAGAAAGG-3′ (anti-sense). The amplification was performed in 35 cycles at 55°C, 1 min; 72°C, 1 min; 94°C, 1 min. After the last cycle, all samples were incubated for an additional 5 min at 72°C. The expression of β-actin was used as an internal control for the assay of a constitutively expressed gene.Real-time PCR was performed with the TaqMan gene expression assay system, using primers and probe mixes for MMP-9 and endogenous GAPDH control genes. PCRs were performed using a 7500 Real-Time PCR System . The primers were: 5′-(TGATGCCATTGCTGATATCCA)-3′ (sense), 5′-(CGGATCCTCAAAGGCTGAGT)-3′ (anti-sense) for MMP-9; 5′-(AACTTTGGCATCGTGGAAGG)-3′ (sense), 5′-(GTGGATGCAGGGATGATGTTC)-3′ (anti-sense) for GAPDH. Relative gene expression was determined by the ΔΔCt method, where Ct meant threshold cycle. All experiments were performed in triplicate (n = 3).Cell lysates containing 1 mg of protein were incubated with 2 μg of anti-TNFR1 antibody at 4°C for 24 h, and then 10 μl of 50% protein A-agarose beads was added and mixed at 4°C for 24 h. The immunoprecipitates were collected and washed thrice with a lysis buffer without Triton X-100. 5X Laemmli buffer was added and subjected to electrophoresis on 12% SDS-PAGE, and then blotted using the anti-TRAF2, anti-c-Src or anti-TNFR1 antibody.A 710-bp (−720 to −11) segment from the 5′-promoter region of the MMP-9 gene was cloned as described . Briefly6 cells/ml (1 ml/well) in 12-well culture plates for 24 h, reaching about 80% confluence. Cells were replaced with 0.4 ml of α-MEM containing 10% FBS. The DNA Metafectene reagent complex was prepared according to the instructions of the manufacturer . The amount of siRNA directed against, ERK2 , JNK2 , p38 , c-Src , TRAF2  or control siRNA was kept at 100 nM for each well. The DNA Metafectene complex (0.1 ml) was added to each well and then incubated at 37°C for 24 h. The cells were washed twice with PBS and maintained in α-MEM containing 1% FBS for 72 h before treatment with TNF-α for the indicated time intervals.MC3T3-E1 cells were plated at 1 × 10MC3T3-E1 cells were seeded in a 10-cm dish. After they reached 90% confluence, cells were starved for 24 h in serum-free α-MEM medium. After stimulation with 15 ng/ml TNF-α for various time intervals, and when inhibitors were used, they were added 1 h prior to the application of TNF-α. As previously described , the celMC3T3-E1 cells were plated on 6-well culture plates with coverslips. Cells were further cultured in serum-free α-MEM for 24 h and treated with 15 ng/ml TNF-α for various time intervals. When inhibitors were used, they were added 1 h prior to the application of TNF-α. After washing twice with ice-cold PBS, the cells were fixed with 4% (w/v) paraformaldehyde in PBS for 30 min, and then permeabilized with 0.3% Triton X-100 in PBS for 15 min. The staining was performed by incubating with 10% normal goat serum in PBS for 30 min followed by incubating with the primary NF-κB (p65) antibody (1:100 dilution) for 1 h in PBS with 1% BSA, washing thrice with PBS, incubating for 1 h with fluorescein isothiocyanate (FITC)-conjugated anti-rabbit antibody (1:100 dilution) in PBS with 1% BSA, washing thrice with PBS, and finally mounting with aqueous mounting medium. The cell nucleus was stained by DAPI. The images were observed under a fluorescence microscope .sICAM-1 released into the culture media of MC3T3-E1 cells was collected and detected by using an ELISA kit according to the manufacturer’s instructions .P < 0.05 was considered significant.Concentration-effect curves were fitted and estimated by using the GraphPad Prism Program . Data were expressed as mean ± S.E.M. and analyzed by one-way ANOVA followed with Tukey’s post-hoc test. TNF-α has been shown to induce the expression of MMP-9 in human osteoblasts, osteoprogenitors, and mesenchymal stem cells ,36. To dTNF receptor l (TNFR1) and TNF receptor-associated factor 2 (TRAF2) generate distinct signals by TNF-α for the induction of differently biological responses in many cell types. Recent evidences suggest that MMP-9 expression was markedly suppressed in TNFR1 KO mice as compared to wild-type mice . PreviouMAPKs, including ERK1/2, p38 MAPK, and JNK1/2, can regulate expression of several genes through activation of downstream kinases or nuclear proteins. Previous study has demonstrated that TNF-α induces MMP-9 expression via p42/p44 MAPK and JNK1/2 in A549 cells . Here, tTo determine whether p38 MAPK is involved in TNF-α-induced MMP-9 expression, a p38 MAPK inhibitor (SB202190) was used. As shown in Figures In addition, to determine whether the activation of JNK1/2 is also involved in TNF-α-induced MMP-9 expression, a pharmacological inhibitor of JNK1/2 SP600125 was used. As shown in Figures Inflammatory responses following stimulation by TNF-α are highly dependent on activation of the transcription factor NF-κB. Moreover, NF-κB is one of the major mediators of the intracellular functions of TNF-α. Therefore, we investigated whether the involvement of NF-κB activation in TNF-α-induced MMP-9 expression in MC3T3-E1 cells, an NF-κB pharmacological inhibitor Bay11-7082 was used. As shown in Figure A significant response was obtained within 5–15 min and declined to the basal level within 30 min. To investigate whether TNF-α stimulates translocation of p65 NF-κB, the cytosolic and nuclear fractions were used to determine the p65 translocation by Western blotting using an anti-p65 antibody. The data showed that TNF-α time-dependently induced translocation of the p65 subunit of NF-κB into nucleus with a significant increase within 15–30 min , MEK1/2 (U0126), p38 MAPK (SB202190), or JNK1/2 (SP600125) for 1 h and then stimulated with TNF-α for the indicated time intervals. Phosphorylation of ERK1/2, p38 MAPK, JNK1/2, IKKα/β and p65 was assayed by Western blotting. As shown in Figure Several studies have shown that up-regulation of MMP-9 mRNA is mediated through an NF-κB-dependent pathway . MMP-9 pPrevious report has shown that TNF-α induces membrane and soluble forms of ICAM-1 release by MMP-9 activity in human osteoblast-like cells . Thus, wi.e. ERK1/2, p38 MAPK, and JNK1/2) and c-Src-independent IKK-NF-κB signaling pathways involved in TNF-α-induced MMP-9 expression and s-ICAM-1 release from MC3T3-E1 cells.MMP-9 is highly expressed in osteoclasts and plays an important role in degradation of ECM. Given the role of osteoblasts in building the bone, it is somewhat counter-intuitive that they also express many matrix degrading MMPs including MMP-2, MMP-3, MMP-8, MMP-9, MMP-13, and MMP-14 . It is gSeveral reports have indicated that most known responses to TNF-α are triggered by binding to one of two distinct receptors, TNFRl and TNFR2, which are differentially regulated on various cell types in normal and diseased tissues. In osteoblasts, TNF-α stimulates osteoblast differentiation through its TNFR1 receptor . Recent Several groups of investigators have reported that TNF-α released during acute and chronic diseases activates multiple intracellular signaling cascades including the MAPKs and NF-κB signaling pathways in various cell types. Previous reports have shown that aggregation of TNFR1/TRAF2 protein complex transducer activates downstream IKKα/β-NF-κB cascade and JNK1/2 and p38 MAPK in skeletal pathologies . TNF-α, i.e. ERK1/2, p38 MAPK, and JNK1/2) and c-Src-independent IKK/NF-κB pathways. MAPKs are serine/threonine protein kinases, which contribute to several cellular pathophysiological responses through regulation of their downstream molecules including transcription factors. Previous studies have indicated that TNF-α induces MMP-9 expression via a MAPK-dependent activation of NF-κB or AP-1 in several cell types [In addition, NF-κB is an inducible transcription factor that plays a key role in the expression of inflammatory response genes. NF-κB plays a pivotal role in bone remodeling cycle . TNF-α bll types ,39,49. HIn bone metabolism, ICAM-1 importantly mediates cell-cell adhesion of osteoblasts and osteoclast precursors, thereby facilitating osteoclast differentiation and bone resorption ,50. OsteOur study provides new insights into the mechanisms by which TNFR1/TRAF2 activates both IKKα/β-NF-κB and c-Src-ERK1/2, p38 MAPK, and JNK1/2 pathways may be associated with MMP-9 expression in osteoblasts-like MC3T3-E1 cells (Figure TNF-α: Tumor necrosis factor-α; MMP-9: Matrix metalloproteinase-9; MC3T3 cells: Mouse osteoblast-like cells; α-MEM: Minimal essential medium-alpha; FBS: Fetal bovine serum; ECL: Enhanced chemiluminescence; BCA: Bicinchoninic acid; PBS: Phosphate-buffered saline; BCA: Bicinchoninic acid; TNFR1: TNF receptor l; TRAF2: TNF receptor-associated factor 2; MAPKs: Mitogen-activated protein kinases; ERK: Extracellular-regulated protein kinase; JNK: c-Jun-N-terminal kinase; sICAM-1: Soluble intercellular adhesion molecule-1; IP: Immunoprecipitation; siRNA: Small interfering RNA; RT-PCR: Reverse transcription-polymerase chain reaction; NF-κB: Nuclear factor-kappa B.The authors declare that they have no competing interests related to this work.Conceived and designed the experiments: CLT, HLH, WCC, and CMY. Acquisition of data: CLT, PLC, and WCC. Analysis and interpretation of data: CMY. Drafting the article: CLT, HLH, WCC and LDH. Revising the article for important intellectual content: HLH and CMY. Final approval of the version to be published: CLT, HLH, PLC, WCC, LDH, and CMY.


Ikkα-floxed mice, and overexpressed IKKα represses carcinogen-induced skin carcinogenesis . We . We IkkαWe have identified similar multi-molecular alterations in mouse and human lung SCCs . Our imm


The transcription factor Helios was constitutively expressed in about one-third of circulating γδ T cells and was upregulated by CD28-signaling. Our data suggest that Helios could serve as a marker for differential activation status rather than for regulatory T cells (Treg). Our findings also indicate that the interaction of CD86 on activated Vδ2 T cells and cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) on activated αβ T cells mediated the suppression because the suppressive effect was abolished by blocking the CD86:CTLA-4 interaction. Pre-treatment of Vδ2 T cells with Toll-like receptor 2 ligands enhanced phosphorylation of MAPKs, Akt, and NF-κB and partially abrogated the suppressive capacity, whereas on co-cultured responder T cells inhibitory molecules were downregulated and Akt and NF-κB phosphorylation was restored. Our results suggest that the regulation of αβ T cell proliferation by activated Vδ2 T cells might contribute to fine-tuning of αβ T cell responses.The proliferation and interleukin-2 production of CD4The online version of this article (doi:10.1007/s00018-013-1467-1) contains supplementary material, which is available to authorized users. In comparison, CD45RA/CD27-negative effector-memory T cells (TEM) and CD45RA-positive and CD27-negative terminally differentiated effector memory T cells (TEMRA) have the ability to home to sites of inflammation where they display immediate effector functions, such as cytokine production and cytotoxicity  as well as in human responder TCRαβ+ FoxP3− T cells . PCH101 targets ther region . Both anSurface expression of CD25 was analyzed on responder T cells, Treg, and γδ T cells after the cells were stained with anti-CD25 mAb clone 2A3 (BD Biosciences).Modulation of inhibitory and co-stimulatory molecules on the surface of responder or γδ T cells after co-culture and before and after stimulation with A/E beads was determined by using the following mAb: anti-CD152 , anti-CD28 (clone L293), anti-PD1 (clone EH12.2H7), anti-PDL-1 (clone MIH1), anti-CD80 (clone L307.4), and anti-CD86 (clone 2331) .5) pre-incubated with medium or a mixture of TLR2L for 22 h followed by a washing step were incubated with 2 μg/mL anti-CD3 (clone OKT3) mAb and with 1 μg/mL anti-CD28 mAb for 30 min on ice. Cells were washed with ice-cold X-VIVO 15 medium, and stimulation was performed by cross-linking of anti-CD3/-CD28 mAb with 10 μg/mL rabbit-anti-mouse (rαm) immunoglobulin Ab for different time periods at 37 °C. Thereafter, cells were fixed with pre-warmed Cytofix™ Buffer  at 37 °C for 10 min, washed, permeabilized with Phosflow™ Perm Buffer III  on ice for 30 min, washed, and stained with phycoerythrin (PE)-conjugated anti-NF-κBp65 (pS529) clone K10-895.12.50, Alexa Fluor 647-labeled mouse anti-phospho-p38 MAPK (pT180/pY182) clone 36/p38, or anti-phospho-Akt-Ser473 clone M89-61 . All samples were analyzed on a FACSCalibur flow cytometer (BD Biosciences) using CellQuestPro software.For the phosphorylation analysis of Akt at S473 and T308, NF-κBp65 at S529, extracellular-signal-regulated kinase (ERK) 1/2 at pT202/pY204, p38 MAPK at pT180/pY182, and Stat3 at Y705, we used the modified Phosflow™ Protocol III (BD Biosciences). Briefly, γδ T cells (2 × 10t test (paired data) was used to analyze the statistical significance of differences.Student’s two-tailed +CD25− responder T cells with CD4+CD25high FoxP3+ Treg stimulated with activation/expander T cell beads (A/E beads) in the absence of APC. As illustrated in Fig. Recently, we reported that activated human Vδ2-expressing γδ T cells negatively regulate the proliferative response of αβ T cells against antigens in the presence of IL-12-producing DC or against strong recall antigens or alloantigens in the presence of APC . To examTo examine whether a subpopulation of Vδ2 γδ T cells exerts this suppressive capacity, we separated Vδ2 γδ T cells according to their CD27 expression into subpopulations of CD27-positive cells  and CD27-negative cells , respectively. In contrast to CD27-negative cells, the majority of the CD27-positive cells co-expressed CD28 (data not shown). Although the suppressive activity of CD27-positive cells was slightly higher than that of CD27-negative cells, we did not observe a significant difference in suppressive function between CD27-negative and CD27-positive cells (data not shown).2O2 scavenger catalase had no effect on the abrogation of suppression (data not shown).In further experiments, we analyzed whether cytotoxic activity of γδ T cells after initial stimulation with A/E beads was responsible for suppression, possibly due to induced cell death of responder T cells. However, we did not observe an increased number of dead cells after co-culturing γδ T cells with responder T cells at the time point when suppression takes place Fig. d. MoreovTaken together, these results indicated that γδ T cells can suppress responder T cells after activation with A/E beads, also in the absence of APC, but that stimulus with anti-CD28 mAb was necessary to induce the suppressive activity of the γδ T cells (data not shown).To obtain more insight into the phenotype of suppressive Vδ2 γδ T cells, we analyzed putative Treg markers such as Helios, FoxP3 (with the clone PCH101 or 259D), and CD25 in/on freshly isolated γδ T cells  and NF-κB in responder T cells. In co-culture with TLR2 ligand pre-treated γδ T cells the inhibition of Akt and NF-κB phosphorylation was abrogated in responder T cells . TLR2 ligand pre-incubation also enhanced Akt and NF-κB phosphorylation in γδ T cells co-cultured with responder T cells, but only slightly in γδ T cells in solo-culture .In summary, the enhanced phosphorylation of MAP kinases, NF-κB, and Akt might provide an explanation for the enhanced production of cytokines and an increased proliferation of responder T cells by γδ T cells after TLR2 ligand treatment.S. aureus enterotoxins), only γδ T cells initially stimulated with A/E beads + TGF-β and IL-15 were still able to suppress the expansion of freshly isolated responder T cells, although FoxP3 was no longer expressed  and, thereafter, cells were re-stimulated before transfer to the patient. We focused on the question of whether BrHPP- or anti-CD3/anti-CD28-activated and -expanded γδ T cells are still able to suppress αβ T cell responses. Moreover, we analyzed if activated and expanded γδ T cells require TGF-β or IL-15 to exert suppressive activity. As shown in Fig. sed Fig. c. Interesed Fig. c. MoreovOur study has identified specific features of suppressive Vδ2 γδ T cells that differentiate them from nTreg. FoxP3 and Helios expression do not represent specific markers for the suppressive capacity of γδ T cells, but the expression of these molecules seems to be involved in the differentiation of γδ T cells with regulatory function. We also provide insights into the suppressive mechanism of freshly isolated Vδ2 γδ T cells. We demonstrate an important role for the interaction of CD86 on γδ T cells and CTLA-4 on responder T cells which induces a reduced phosphorylation of Akt and NF-κB. Furthermore, we show that the suppressive mechanism of γδ T cells was abrogated by TLR2 ligands, inducing a strong Th1 response and abolishing the γδ T cell-mediated inhibition of Akt and NF-κB phosphorylation in responder T cells .+ responder T cells. The expression of FoxP3 detected with PCH101 mAb but not with 259D mAb has been demonstrated in responder T cells, but it did not coincide with the regulatory function of these cells [S. aureus enterotoxins) can be due to an upregulation of intracellular GATA-3 in these γδ T cells. The transcription factor GATA-3 controls the production of immunoregulatory cytokines that could specifically counteract a Th1 response. Hansmann et al. [CM γδ T cells produce less IFN-γ than TEM γδ T cells. We observed that previously FoxP3-expressing Vδ2 γδ T cells, which were still suppressive after initial stimulation with A/E beads and TGF-β/IL-15/IL-2, co-expressed CD27 and Helios, whereas CD27 expression decreased in Helios-negative previously FoxP3-expressing Vδ2 γδ T cells after stimulation with BrHPP and TGF-β/IL-15/IL-2 . In this context, conflicting data exist on the function of Helios. Thornton and co-workers [Natural Treg, which regulate peripheral tolerance, highly express CTLA-4 and CD25 and can be stained to the same extent with FoxP3 mAb 259D and PCH101 . In our se cells . In addise cells , 50–52. se cells , we obsese cells , we demon et al.  demonstrn et al.  demonstr-workers  describe-workers –30. Simi-workers , we obse-workers . Our res-workers . Studies-workers  on the g-workers –58. In t-workers . Further-workers  who demoγδ T cells display a broad range of functional plasticity after activation, and Vδ2 γδ T cells can be differentiated on the basis of polarizing cytokines, homing receptors, and lineage-determining transcription factors. Several recent publications have reported that Vδ2 γδ T cells can express an APC-like phenotype after pre-activation for 18–48 h with PAg, autologous B cells, or irradiated HLA-A2-negative Epstein–Barr virus-transformed B cell lines in the presence of IL-2 and IL-15, characterized by an upregulation of APC-specific molecules and APC function , 60. AddWhile initially A/E beads-stimulated γδ T cells were suppressive, pre-treatment with TLR2 ligands partially abolished suppression and led to an enhanced phosphorylation of MAPKs, Akt, and NF-κB resulting in a higher production of Th1 cytokines. We and others have already described that anti-CD3 mAb as well as PAg activation of γδ T cells induce phosphorylation of MAPK ERK2 and of p38 and Akt (at serine 473), which in turn induce the upregulation of crucial target genes, such as IFN-γ and TNF-α –65. MoreS. aureus enterotoxins that activate a greater Th1-repertoire of responder T cells than only one S. aureus enterotoxin . It is  T cells . We obse T cells . Enhance T cells , 69. TheIn conclusion, our data provide evidence that Vδ2 γδ T cells have an immunosuppressive potential in the presence of APC  or afterSupplemental Fig. 1 Suppression is not mediated by IL-2 competition, but is cell–cell contact dependent. a 104 responder T cells  were cultured alone or co-cultured with 104 γδ T cells in the absence or presence of exogenous IL-2 (50 U/mL) for 7 days. After activation with A/E beads, the absolute cell numbers of viable responder T cells and γδ T cells were determined 7 days after culture by SCDA. The mean value of quadruplicate  or triplicate (γδ solo-culture + IL-2) measurement for each donor is shown as one symbol. The black bars represent the mean values of 4 different experiments. b 104 responder T cells (Resp) were cultured alone or co-cultured with 104 γδ T cells separated from them by the membrane of a trans-well insert. Resp and γδ T cells were stimulated by A/E beads and cultivated for 7 days. Absolute cell numbers of viable responder T cells was determined by SCDA (TIFF 562 kb)Supplemental Fig. 2 Influence of TLR2-ligands on the cytokine production. 104 responder T cells were cultured in medium alone or with 104 γδ T cells untreated or pre-treated with a mixture of TLR2 ligands as indicated. After stimulation with A/E beads, supernatants were collected and analyzed by ELISA following the instructions of the manufacturer . IFN-γ, TNF-α, and granzyme B production was determined after 72 h and IL-2 or RANTES after 96 h. Each symbol represents one donor, and the bars represent the mean value of different experiments as indicated. Asterisks indicate statistical significance (*p ≤ 0.05 and **p ≤ 0.01) (TIFF 717 kb)Supplemental Fig. 3 Phosphorylation of signaling molecules in γδ T cells is enhanced after TLR2-L pre-treatment. 106 γδ T cells were pre-treated in medium or with a mixture of TLR2-L and thereafter stimulated with anti-CD2, anti-CD3, and anti-CD28 mAb cross-linked with 10 µg/mL rαm Ig for the indicated time points. a Cells were lysed in NP40 lysis buffer  with 1% (v/v) of detergent in 20 mM Tris-HCl, 150 mM NaCl with protease inhibitors aprotinin, leupeptin, PMSF, sodium fluoride, and sodium pyrophosphate. Samples were separated on 10% SDS-gel, and protein was transferred to nitrocellulose membranes . Blots were blocked with 5% BSA and phosphorylated molecules were detected by protein phosphorylation-specific antibodies as indicated. As a loading control, blots were stripped and reprobed with antibodies (as indicated) detecting whole protein levels or with anti-β-actin mAb. Primary Abs were detected by the appropriate HRP-conjugated antibody . Numbers represent densitometric evaluation. b The bars present the values of densitometric evaluation in relation to the corresponding control (TIFF 1099 kb)Supplemental Fig. 4 Phosphorylation of signalling molecules in γδ T cells co-cultured with responder T cells. 104 responder T cells were cultured alone or in the presence of freshly isolated γδ T cells pre-treated in medium or with a TLR2-L mixture. After A/E bead stimulation, cells were cultured for 3 days, then fixed and subsequently permeabilized. Indicated phosphorylated signaling molecules were labeled with specific fluochrome-conjugated antibodies and analyzed by flow cytometry. Mean values of the median fluorescence intensity of at least 4 donors is shown. Each symbol represents the data of one donor, and the black bars present the mean value for 4 different experiments. Asterisks indicate statistical significance . n.s. Non-significant) (TIFF 459 kb)Supplemental Fig. 5 Helios is induced after co-stimulation with anti-CD28 mAb. Co-expression of CD27 and Helios was stained with the indicated antibodies. The expression was analyzed by flow cytometry on/in γδ T cells after stimulation with anti-CD3 mAb or A/E beads (anti-CD3 and anti-CD28 mAb coated) after 8 days of cell culture (TIFF 300 kb)Below is the link to the electronic supplementary material.