Immunologic and clinical improvement of progressive coccidioidomycosis following administration of transfer factor

Three patients with progressive coccidioidomycosis were given preparations of transfer factor (TF). Adverse reactions to TF were minimal. Following TF administration two of these patients had prolonged clinical remissions in their coccidioidal disease. Cellular immune responses were sequentially evaluated by coccidioidininduced delayed-type skin tests, lymphocyte blast transformation and macrophage inhibition factor production (MIF). These three patients each exhibited different cellular immune patterns before and after TF administration. Two patients converted their coccidioidin skin tests, and one converted lymphocyte transformation response to coccidioidin. Also, TF apparently favorably affected the MIF response in all three patients.     

Structural changes in thin filaments of crab striated muscle.

Low angle X-ray diffraction patterns were recorded from crab leg muscle in living resting state and in rigor (glycerol-extracted). Both resting and rigor patterns showed a series of layer-lines arising from a helical arrangement of actin subunits in the thin filaments. In the resting state, the crossover repeat of the long-pitch actin helices was 36.6 nm, and the symmetry of the genetic actin helix was an intermediate between $\text{26}\text{12}$ and $\text{28}\text{13}$. When the muscle went into rigor, the crossover repeat changed to 38.3 nm and the helical symmetry to $\text{28}\text{13}$.In the living resting pattern, six other reflections were observed on the meridian and in the near-meridional region. These were indexed as orders of 2 × 38.2 nm and could be assigned to troponin molecules; the spacings and the intensity distributions of these reflections could be explained by the model proposed by Ohtsuki (1974) for the arrangement of troponin molecules in the thin filaments.The muscle in rigor gave meridional and near-meridional reflections at orders of 2 × 38.3 nm. These were identified as the same series of reflections as was assigned to troponin in the living resting pattern, but were more intense and could be seen up to higher orders. We consider that the myosin heads attached to the thin filament at regular intervals along its axis also contribute to these reflections in the rigor pattern.     

Actin filaments in muscle: Pattern of myosin and tropomyosin/troponin attachments

X-ray patterns from lobster and crayfish muscles show very clear layer lines from the thin filaments, well separated from the myosin layer lines. The intensities in patterns from relaxed muscles include an important contribution from the regulatory proteins, and allow the arrangement of the troponin complexes to be deduced. Moreover, the troponin diffraction indirectly provides an accurate value for the pitch of the actin helix in relaxed muscle.In rigor, the attachment of cross-bridges modifies the intensities. These X-ray patterns support Reedy's (1968) concept that cross-bridges in rigor attach only to certain azimuths on the actin filaments (“target areas”); the 145 Å repeat of their origins on the thick filaments is not reflected in the pattern of attachment. Our calculations show that the observed intensities agree quantitatively with those expected for models based on such attachment, but depend significantly on the locations of the troponin complexes. The arrangement of the filament components is discussed in terms of design requirements. Our conclusions may be applicable to many other muscles, especially insect flight muscle and other invertebrate muscles.     

Anion and ionic strength effects upon the oxidation of cytochrome c by cytochrome c oxidase

Citrate and other polyanion binding to ferricytochrome c partially blocks reduction by ascorbate, but at constant ionic strength the citrate-cytochrome c complex remains reducible; reduction by TMPD is unaffected. At a constant high ionic strength citrate inhibits the cytochrome c oxidase reaction competitively with respect to cytochrome c, indicating that ferrocytochrome c also binds citrate, and that the citrateferrocytochrome c complex is rejected by the binding site at high ionic strength. At lower ionic strengths, citrate and other polyanions change the kinetic pattern of ferrocytochrome c oxidation from first-order towards zero-order, indicating preferential binding of the ferric species, followed by its exclusion from the binding site. The turnover at low cytochrome c concentrations is diminished by citrate but not the K_m (apparent non-competitive inhibition) or the rate of cytochrome a reduction by bound cytochrome c. Small effects of anions are seen in direct measurements of binding to the primary site on the enzyme, and larger effects upon secondary site binding. It is concluded that anion-cytochrome c complexes may be catalytically competent but that the redox potentials and/or intramolecular behaviour of such complexes may be affected when enzyme-bound. Increasing ionic strength diminishes cytochrome c binding not only by decreasing the ‘association’ rate but also by increasing the ‘dissociation’ rate for bound cytochrome c converting the ‘primary’ (T) site at high salt concentrations into a site similar kinetically to the ‘secondary’ (L) site at low ionic strength. A finite K_m of 170 μM at very high ionic strength indicates a ratio of $\text{K}{}^{\mathrm{\infty }}{}_{\mathrm{<mtext>M</mtext>}}\text{K}{}^0{}_{\mathrm{<mtext>M</mtext>}}$ of about 5000. It is proposed that anions either modify the E¹₀ of cytochrome c bound at the primary (T) site or that they perturb an equilibrium between two forms of bound c in favour of a less active form.     

宁夏东部风沙区固定沙丘土壤性质小尺度空间变异特征

随着流动沙丘的不断固定,不同地形部位的土壤机械组成、有机碳(SOC)和全氮(TN)含量的变化存在差异。为了探明固定沙丘的土壤机械组成与土壤养分空间变异特征,本研究以宁夏黄沙古渡固定沙丘为对象,分析了固定沙丘不同部位、不同土层的风沙土机械组成、SOC和TN特征。结果表明: 各土层土壤机械组成均以中沙和细沙为主。SOC和TN有明显的表聚作用,SOC和TN含量最大值分别为5.781和0.412 g·kg^-1,分布在丘间地,脊线和背风坡含量最少。随着土层深度的增加,背风坡和迎风坡的SOC含量逐渐减少,丘间地的SOC含量先减少后增加。SOC和TN在小尺度上均表现出明显的空间异质性;SOC和TN与粉沙和极细沙含量呈显著正相关,与中沙和粗沙含量呈显著负相关。土壤机械组成对SOC和TN含量有明显影响,而且SOC和TN含量随着细颗粒含量的增加而增加,表明土壤细颗粒对有机质的吸附和积累起着重要作用。     

SAR inspired by aldehyde oxidase (AO) metabolism: Discovery of novel,CNS penetrant tricyclic M4 PAMs

This letter describes progress towards an M₄ PAM preclinical candidate inspired by an unexpected aldehyde oxidase (AO) metabolite of a novel, CNS penetrant thieno[2,3-c]pyridine core to an equipotent, non-CNS penetrant thieno[2,3-c]pyrdin-7(6H)-one core. Medicinal chemistry design efforts yielded two novel tricyclic cores that enhanced M₄ PAM potency, regained CNS penetration, displayed favorable DMPK properties and afforded robust in vivo efficacy in reversing amphetamine-induced hyperlocomotion in rats.     

The PDZ-GEF Gef26 regulates synapse development and function via FasII and Rap1 at the Drosophila neuromuscular junction

Guanine nucleotide exchange factors (GEFs) are essential for small G proteins to activate their downstream signaling pathways, which are involved in morphogenesis, cell adhesion, and migration. Mutants of Gef26, a PDZ-GEF (PDZ domain-containing guanine nucleotide exchange factor) in Drosophila, exhibit strong defects in wings, eyes, and the reproductive and nervous systems. However, the precise roles of Gef26 in development remain unclear. In the present study, we analyzed the role of Gef26 in synaptic development and function. We identified significant decreases in bouton number and branch length at larval neuromuscular junctions (NMJs) in Gef26 mutants, and these defects were fully rescued by restoring Gef26 expression, indicating that Gef26 plays an important role in NMJ morphogenesis. In addition to the observed defects in NMJ morphology, electrophysiological analyses revealed functional defects at NMJs, and locomotor deficiency appeared in Gef26 mutant larvae. Furthermore, Gef26 regulated NMJ morphogenesis by regulating the level of synaptic Fasciclin II (FasII), a well-studied cell adhesion molecule that functions in NMJ development and remodeling. Finally, our data demonstrate that Gef26-specific small G protein Rap1 worked downstream of Gef26 to regulate the level of FasII at NMJs, possibly through a βPS integrin-mediated signaling pathway. Taken together, our findings define a novel role of Gef26 in regulating NMJ development and function.     

A single nuclear polymorphism in let-7g binding site affects the doubling time of thyroid nodule by regulating KRAS-induced cell proliferation

As an indicator for the malignancy of thyroid nodules (TN), the doubling time of TN was studied in this study to evaluate the effect of rs712 polymorphism on the progression of TN. In addition, we aimed to study the potential molecular mechanisms underlying the pathological effect of rs712 polymorphism upon TN. A Taqman method was used to genotype the patients according to their rs712 polymorphism. Real-time polymerase chain reaction, western blot, Terminal deoxynucleotidyl transferase dUTP nick end labeling assay and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium (MTT) assay was conducted to study the correlation between KRAS expression and the pathological effect of rs712 polymorphism. In-silicon analysis and luciferase assay were utilized to establish the regulatory relationship between let-7g and KRAS. KRAS messenger RNA (mRNA)/protein levels in the GG group were upregulated with a decreased apoptosis index. KRAS mRNA was validated to be a virtual target of let-7g. In addition, the mRNA/protein level of KRAS as well as cell proliferation index was decreased in primary thyroid cancer cells genotyped as TT/TG and transfected with KRAS small interfering RNA (siRNA)/let-7g precursors. The cell apoptosis index was evidently elevated in the KRAS siRNA/let-7g precursors group compared with that in the scramble controls. Moreover, KRAS mRNA/protein only showed slight reduction when GG-genotyped primary thyroid cancer cells were transfected by let-7g precursors. Additionally, let-7g precursors exhibited no significant effect on cell proliferation index or cell apoptosis in GG cells. Rs712 polymorphism T>G in the 3′-untranslated region of KRAS interrupts the interactions between let-7g and KRAS mRNA, leading to a higher cell proliferation index and reduced doubling time of TN.     

The influence of fundamental design parameters on ciliates community structure in Irish activated sludge systems

The protozoan community in eleven activated sludge wastewater treatment plants (WWTPs) in the greater Dublin area has been investigated and correlated with key physio-chemical operational and effluent quality parameters. The plants represented various designs, including conventional and biological nutrient removal (BNR) systems. The aim of the study was to identify differences in ciliate community due to key design parameters including anoxic/anaerobic stages and to identify suitable bioindicator species for performance evaluation. BNR systems supported significantly different protozoan communities compared to conventional systems. Total protozoan abundance was reduced in plants with incorporated anoxic and anaerobic stages, whereas species diversity was either unaffected or increased. Plagiocampa rouxi and Holophrya discolor were tolerant to anoxic/anaerobic conditions and associated with high denitrification. Apart from process design, influent wastewater characteristics affect protozoan community structure. Aspidisca cicada was associated with low dissolved oxygen and low nitrate concentrations, while Trochilia minuta was indicative of good nitrifying conditions and good sludge settleability. Trithigmostoma cucullulus was sensitive to ammonia and phosphate and could be useful as an indicator of high effluent quality. The association rating assessment procedure of Curds and Cockburn failed to predict final effluent biological oxygen demand (BOD₅) indicating the method might not be applicable to treatment systems of different designs.