Maximum likelihood estimation in the additive hazards model

The additive hazards model specifies the effect of covariates on the hazard in an additive way, in contrast to the popular Cox model, in which it is multiplicative. As the non-parametric model, additive hazards offer a very flexible way of modeling time-varying covariate effects. It is most commonly estimated by ordinary least squares. In this paper, we consider the case where covariates are bounded, and derive the maximum likelihood estimator under the constraint that the hazard is non-negative for all covariate values in their domain. We show that the maximum likelihood estimator may be obtained by separately maximizing the log-likelihood contribution of each event time point, and we show that the maximizing problem is equivalent to fitting a series of Poisson regression models with an identity link under non-negativity constraints. We derive an analytic solution to the maximum likelihood estimator. We contrast the maximum likelihood estimator with the ordinary least-squares estimator in a simulation study and show that the maximum likelihood estimator has smaller mean squared error than the ordinary least-squares estimator. An illustration with data on patients with carcinoma of the oropharynx is provided.     

The Effect of Z-Ligustilide on the Mobility of Human Glioblastoma T98G Cells

Z-ligustilide (LIG), an essential oil extract from Radix Angelica sinensis, has broad pharmaceutical applications in treating cardio-vascular diseases and ischemic brain injury. Recently, LIG has been connected to Glioblastoma multiforme (GBM) because of its structural similarity to 3-n-alkyphthalide (NBP), which is specifically cytotoxic to GBM cells. Hence, we investigated LIG’s effect on GBM T98G cells. The study shows that LIG can significantly reduce T98G cells’ migration in a dose-dependent manner. Furthermore, the attenuation of cellular mobility can be linked to the activity of the Rho GTPases (RhoA, Rac1 and Cdc42), the three critical molecular switches governing cytoskeleton remodeling; thus, regulating cell migration. LIG significantly reduces the expression of RhoA and affects in a milder manner the expression of Cdc42 and Rac1.     

A stable method for Agrobacterium-mediated transformation of Mentha piperita and Mentha canadensis using internodal explants

In Vitro Cellular & Developmental Biology - Plant - Mentha species are planted worldwide for extraction of their metabolites, including essential oils and phenolic acid compounds, which have...     

Atypical OmpR/PhoB Subfamily Response Regulator GlnR of Actinomycetes Functions as a Homodimer,Stabilized by the Unphosphorylated Conserved Asp-focused Charge Interactions

The OmpR/PhoB subfamily protein GlnR of actinomycetes is an orphan response regulator that globally coordinates the expression of genes related to nitrogen metabolism. Biochemical and genetic analyses reveal that the functional GlnR from Amycolatopsis mediterranei is unphosphorylated at the potential phosphorylation Asp⁵⁰ residue in the N-terminal receiver domain. The crystal structure of this receiver domain demonstrates that it forms a homodimer through the α4-β5-α5 dimer interface highly similar to the phosphorylated typical response regulator, whereas the so-called “phosphorylation pocket” is not conserved, with its space being occupied by an Arg⁵² from the β3-α3 loop. Both in vitro and in vivo experiments confirm that GlnR forms a functional homodimer via its receiver domain and suggest that the charge interactions of Asp⁵⁰ with the highly conserved Arg⁵² and Thr⁹ in the receiver domain may be crucial in maintaining the proper conformation for homodimerization, as also supported by molecular dynamics simulations of the wild type GlnR versus the deficient mutant GlnR(D50A). This model is backed by the distinct phenotypes of the total deficient GlnR(R52A/T9A) double mutant versus the single mutants of GlnR (i.e. D50N, D50E, R52A and T9A), which have only minor effects upon both dimerization and physiological function of GlnR in vivo, albeit their DNA binding ability is weakened compared with that of the wild type. By integrating the supportive data of GlnRs from the model Streptomyces coelicolor and the pathogenic Mycobacterium tuberculosis, we conclude that the actinomycete GlnR is atypical with respect to its unphosphorylated conserved Asp residue being involved in the critical Arg/Asp/Thr charge interactions, which is essential for maintaining the biologically active homodimer conformation.     

Heteromerization of short-chain trans-prenyltransferase controls precursor allocation within a plastidial terpenoid network

Terpenes are the largest and most diverse class of plant specialized metabolites. Sesterterpenes(C25), which are derived from the plastid methylerythritol phosphate pathway,were recently characterized in plants. In Arabidopsis thaliana, four genes encoding geranylfarnesyl diphosphate synthase(GFPPS)(AtGFPPS1 to 4) are responsible for the production of GFPP, which is the common precursor for sesterterpene biosynthesis. However,the interplay between sesterterpenes and other known terpenes remain e...     

Purification,characterization, and gene cloning of two laccase isoenzymes (Lac1 and Lac2) from Trametes hirsuta MX2 and their potential in dye decolorization

In this study, two laccase isoenzymes (Lac1 and Lac2) from the culture supernatant of Trametes hirsuta MX2 were purified, and the genes (Lac1 and Lac2) coding the isoenzymes were cloned. Both Lac1 and Lac2 contained an open reading frame of 1563 bp with an identity of 79%. The two isoenzymes showed significant biochemical differences. The maximal activities of Lac1 and Lac2 were at pH 2.5 with 2-2′-azino-di-(3-ethylbenzthiazoline sulfonic acid) (ABTS), and the optimal temperatures for the activities of Lac1 and Lac2 were 60 and 50 °C, respectively. Lac1 exhibited excellent resistance to acidic conditions and retained 62.17% of its initial activity at pH 2.5 after a 72-h incubation. Lac2 was more thermostable than Lac1 with half-lives (t_1/2) of 9.58 and 3.12 h at 50 and 60 °C, respectively; the t_1/2 of Lac1 were only 4.19 and 0.88 h, respectively. Both Lac1 and Lac2 isoenzymes have a strong tolerance to Mg²⁺, Mn²⁺, Cu²⁺, and EDTA (50 mM). At a low concentration of 0.05 U mL^?1, the enzymes could decolorize towards Remazol Brilliant Blue R, Acid Red 1, Crystal Violet, and Neutral Red in the presence of ABTS. These unusual properties demonstrated that the two laccases have strong potential for specific industrial applications.

     

Reciprocal regulation between ER stress and autophagy in renal tubular fibrosis and apoptosis

Both endoplasmic reticulum (ER) stress and autophagy have been implicated in chronic kidney injury and renal fibrosis. However, the relationship and regulatory mechanisms between ER stress and autophagy under this condition remain largely unknown. In this study, we first established a mouse model of ER stress-induced chronic kidney injury by 2 weekly injections of a low dose of tunicamycin (TM), a classical ER stress inducer. This model showed the induction of ER stress, autophagy, fibrosis and apoptosis in kidney tissues. In vitro, TM also induced ER stress, autophagy, fibrosis and apoptosis in HK-2 human kidney proximal tubular cells and BUMPT-306 mouse kidney proximal tubular cells. In these cells, autophagy inhibitor suppressed TM-induced fibrotic changes and apoptosis, suggesting an involvement of autophagy in ER stress-associated chronic kidney injury. PERK inhibitor ameliorated autophagy, fibrotic protein expression and apoptosis in TM-treated cells, indicating a role of the PERK/eIF2α pathway in autophagy activation during ER stress. Similar results were shown in TGF-β1-treated HK-2 cells. Interestingly, in both TM- or TGF-β1-treated kidney proximal tubular cells, inhibition of autophagy exaggerated ER stress, suggesting that autophagy induced by ER stress provides a negative feedback mechanism to reduce the stress. Together, these results unveil a reciprocal regulation between ER stress and autophagy in chronic kidney injury and fibrosis.Subject terms: Acute kidney injury, Chronic kidney disease     

CADASIL mutant NOTCH3(R90C) decreases the viability of HS683 oligodendrocytes via apoptosis

Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is the most common hereditary cerebral small vessel disease caused by mutations in NOTCH3. Prevailing models suggest that demyelination occurs secondary to vascular pathology. However, in zebrafish, NOTCH3 is also expressed in mature oligodendrocytes. Thus, we hypothesized that in addition to vascular defects, mutant NOTCH3 may alter glial function in individuals with CADASIL. The aim of this study was to characterize the direct effects of a mutant NOTCH3 protein in HS683 oligodendrocytes. HS683 oligodendrocytes transfected with wild-type NOTCH3, mutant NOTCH3(R90C), and empty control vector were used to study the impact of the NOTCH3(R90C) mutant on its protein hydrolytic processing, cell viability, apoptosis, autophagy, oxidative stress, and the related upstream events using immunoblotting, immunofluorescence, RT-PCR, and flow cytometry. We determined that HS683 oligodendrocytes transfected with mutant NOTCH3(R90C), which is the hotspot mutation site-associated with CADASIL, exhibited aberrant NOTCH3 proteolytic processing. Compared to cells overexpressing wild-type NOTCH3, cells overexpressing NOTCH3(R90C) were less viable and had a higher rate of apoptosis. Immunoblotting revealed that cells transfected with NOTCH3(R90C) had higher levels of intrinsic mitochondrial apoptosis, extrinsic death receptor path-related apoptosis, and autophagy compared with cells transfected with wild-type NOTCH3. This study suggests that in patients with CADASIL, early defects in glia influenced by NOTCH3(R90C) may directly contribute to white matter pathology in addition to secondary vascular defects. This study provides a potential therapeutic target for the future treatment of CADASIL.