Activation of a non-specific cation conductance by intracellular injection of inositol 1,3,4,5-tetrakisphosphate into identified neurons of Aplysia

The ionic mechanism of the effect of intracellulary injected inositol 1,3,4,5-tetrakisphosphate (IP₄) on the membrane of identified neurons (R9–R12) of Aplysia kurodai was investigated with conventional voltage-clamp, pressure injection, and ion-substitution techniques. Intracellular injection of IP₄ into a neuron voltage-clamped at −45 mV reproducibly induced a slow inward current (20–60 s in duration, 3–5 nA in amplitude) associated with a conductance increase. The current was decreased by depolarization and increased by hyperpolarization. The extrapolated reversal potential was −21 mV. The IP₄-induced inward current was sensitive to changes in the external Na⁺, Ca²⁺ and K⁺ concentration but not to changes in Cl⁻ concentration, and was resistant to tetrodotoxin (50 μM). When the cell was perfused with tetraethylammonium (5 mM) but not with 4-aminopyridine (5 mM), the IP₄-induced inward current recorded at −45 mV slightly increased. The IP₄-induced inward current was partially reduced by calcium channel blockers (Co²⁺ and Mn²⁺). These results suggest that intracellularly injected IP₄ can activate a non-specific cation conductance.     

Invasive pulmonary aspergillosis in a puerperant with drug-induced agranulocytosis.

Invasive pulmonary aspergillosis (IPA) is an acute infection of Aspergillus species to the lungs. It generally occurs in immunocompromised hosts, especially with neutropenia. We report a 30-year-old puerperant, who developed IPA from agranulocytosis. She had been treated for threatened labor with ritodrine and cefepime, one of which induced agranulocytosis. After vaginal delivery of twins, pneumonia emerged in the right lower lobe. She was diagnosed to have IPA according to the halo sign on computed tomography (CT) and positive circulating antibody against Aspergillus, and was treated successfully with oral itraconazole followed by surgical resection. It is important to note that IPA might arise in otherwise immunocompetent hosts when neutropenia is long-standing.     

Drug-induced liver injury caused by an herbal medicine, bofu-tsu-sho-san

A 37-year-old woman was admitted to a hospital with jaundice. Within a couple of weeks, her liver function improved with only symptomatic therapy. About 30 to 60 days before admission, she had taken a herbal medicine, bofu-tsu-sho-san. A diagnosis of drug-induced liver injury was made according to the diagnostic scale proposed at the Digestive Disease Week-Japan 2004. A drug-lymphocyte stimulation test for each ingredient of bofu-tsu-sho-san; the results were positive for Cnidii Rhizoma, Angelicae Radix and Menthae Herba. The liver biopsy specimen revealed features of acute hepatitis. Physicians should be aware that bofu-tsu-sho-san can cause liver injury, as this drug is commonly used as an over-the-counter medicine.     

Decline in estimated glomerular filtration rate is associated with risk of end-stage renal disease in type 2 diabetes with macroalbuminuria: an observational study from JDNCS

Background

There is increased interest in surrogate endpoints for clinical trials of chronic kidney disease.

Methods

In this nationwide observational study of 456 patients with type 2 diabetes and clinically suspected diabetic nephropathy followed for a median of 4.2 years, we evaluated the association between estimated glomerular filtration rate (eGFR) and albuminuria at baseline or during follow-up and risk of ESRD.

Results

Low eGFR (<60 mL/min/1.73 m²) and macroalbuminuria at enrollment were independently associated with risk of ESRD. In patients with macroalbuminuria, both ≤?50% change and ?50 to ?30% change in eGFR over 1 and 2 years were predictive of ESRD. The higher cut point (≥50% decline in eGFR) was more strongly predictive but less common. Remission of macroalbuminuria to normo-/microalbuminuria at 1 and 2 years was associated with a lower incidence of ESRD than no remission; however, it was not a determinant for ESRD independently of initial eGFR and initial protein-to-creatinine ratio.

Conclusion

These results suggest that a ≥30% decline in eGFR over 1 or 2 years adds prognostic information about risk for ESRD in patients with type 2 diabetes and macroalbuminuria, supporting the consideration of percentage decline in eGFR as a surrogate endpoint among macroalbuminuric cases in type 2 diabetes. On the other hand, our study suggests that additional analyses on the relationship between remission of macroalbuminuria and risk of ESRD are needed in type 2 diabetes.

     

Enriched environments influence depression-related behavior in adult mice and the survival of newborn cells in their hippocampi

Major depression is a highly prevalent mental disorder and environmental factors have been strongly implicated in its pathophysiology. Clinical studies have demonstrated that stress or depression can lead to atrophy and cell loss in the hippocampus. Studies of animal models of depression have suggested that reduced neurogenesis in the adult hippocampus might contribute to such structural changes and to the behavior of these animals. On the other hand, increased hippocampal neurogenesis can be induced by the administration of antidepressants or electroconvulsive seizure, suggesting that increased neurogenesis might be related to the treatment of depression. Thus, an enriched environment (EE), which also enhances neurogenesis, is expected to have therapeutic effects on depression-related behaviors. To investigate the effects of an EE during adulthood on these behaviors, we subjected adult mice housed in an EE for five weeks to behavioral tests. In an open field test, EE mice exhibited a decrease in the distance traveled and an increase in the amount of time spent in the center. The startle response was smaller in EE mice than in control mice. EE mice also showed reduced immobility time in a forced swim test. The immobility time in EE mice was approximately half that observed in mice treated with a tricyclic antidepressant, imipramine. In our experimental condition, increased survival of newborn cells was observed in EE mice by 5-bromo-2'-deoxyuridine (BrdU)-labeled immunohistochemistry. Double-staining of BrdU and a mature neuron marker, NeuN, revealed that the majority of surviving cells were neurons. Our results suggest that EE, which enhanced the survival of newborn neurons, shows beneficial effects on behavioral despair and habituation to a novel environment.     

Overexpression of gamma-glutamyltransferase in transgenic mice accelerates bone resorption and causes osteoporosis

We previously identified gamma-glutamyltransferase (GGT) by expression cloning as a factor inducing osteoclast formation in vitro. To examine its pathogenic role in vivo, we generated transgenic mice that overexpressed GGT in a tissue-specific manner utilizing the Cre-loxP recombination system. Systemic as well as local production of GGT accelerated osteoclast development and bone resorption in vivo by increasing the sensitivity of bone marrow macrophages to receptor activator of nuclear factor-kappaB ligand, an essential cytokine for osteoclastogenesis. Mutated GGT devoid of enzyme activity was as potent as the wild-type molecule in inducing osteoclast formation, suggesting that GGT acts not as an enzyme but as a cytokine. Recombinant GGT protein increased receptor activator of nuclear factor-kappaB ligand expression in marrow stromal cells and also stimulated osteoclastogenesis from bone marrow macrophages at lower concentrations. Thus, GGT is implicated as being involved in diseases characterized by accelerated osteoclast development and bone destruction and provides a new target for therapeutic intervention.     

IL-17A stimulates the expression of inflammatory cytokines via celecoxib-blocked prostaglandin in MC3T3-E1 cells

Objective

The prostaglandins (PGs) released from osteoblasts can alter the process of bone remodelling. Recently, we showed that compressive force induced the expression of pro-inflammatory cytokine interleukin (IL)-17s and their receptors in osteoblastic MC3T3-E1 cells and that IL-17A was expressed most highly. Consequently, in the current study we examined the effect of IL-17A and/or celecoxib on PGE₂ production and the expression of cyclooxygenases (COXs) and inflammatory cytokines in MC3T3-E1 cells. We also examined the effects of PGE₂ and cyclohexamide on the expression of inflammatory cytokines.

Methods

Cells were cultured with or without IL-17A (0.1, 1.0, or 10 ng/ml) in the presence or absence of 10 μM celecoxib, a specific inhibitor of COX-2, for up to 72 h. Cells were pretreated with or without 10 μg/ml cycloheximide, protein synthesis inhibitor, for 30 min, and then cultured with 10 ng/ml IL-17A for 24 h. Cells were also cultured with or without 1.5 ng/ml PGE₂ for 24 h. PGE₂ production was determined by ELISA. The expression of COX-1, COX-2, IL-1α, IL-6, IL-8, IL-11, and TNF-α mRNAs and proteins was determined by real-time PCR and ELISA, respectively.

Results

The expression of COX-2, IL-1α, IL-6, IL-8, IL-11, and TNF-α, as well as PGE₂ production increased in the presence of IL-17A, whereas COX-1 expression did not change. Celecoxib blocked the stimulatory effect of IL-17A on the expression of COX-2, IL-1α, IL-6, IL-8, and IL-11 as well as PGE₂ production, whereas it did not block TNF-α expression. Cycloheximide pretreatment suppressed the expression of IL-17-induced inflammatory cytokines. The expression of IL-1α, IL-6, IL-8, and IL-11 increased by the addition of PGE₂, whereas TNF-α expression was not affected.

Conclusion

These results suggest that IL-17A stimulates the expression of bone resorption-related inflammatory cytokines through an autocrine mechanism involving celecoxib-blocked PGs, mainly PGE₂, in osteoblasts.     

Developmental and spatial expression pattern of alpha-taxilin in the rat central nervous system

Alpha-taxilin has been identified as a binding partner of syntaxin family members and thus has been proposed to function in syntaxin-mediated intracellular vesicle trafficking. However, the lack of detailed information concerning the cellular and subcellular localization of alpha-taxilin impedes an understanding of the role of this protein. In the present study, we characterized alpha-taxilin-expressing cells in the rat CNS with a specific antibody. During embryonic development, alpha-taxilin was prominently expressed in nestin-positive neural stem cells in vivo and in vitro. As CNS development proceeded, the alpha-taxilin expression level was rapidly down-regulated. In the postnatal CNS, alpha-taxilin expression was almost confined to the neuronal lineage, with the highest levels of expression in motor neurons within the brainstem nuclei and spinal cord and in primary sensory neurons in mesencephalic trigeminal nucleus. At the cellular level, alpha-taxilin was preferentially located in Nissl substance-like structures with a tigroid or globular morphology within the soma and proximal to dendrites, but it was excluded from terminals. Combined staining with propidium iodide demonstrated that alpha-taxilin distribution overlapped with the cytoplasmic compartment enriched in RNA species, suggesting a close association of alpha-taxilin with actively translating ribosomes or polysomes in neurons. In agreement with this, a recent study indicated the preferential binding of alpha-taxilin to the nascent polypeptide-associated complex (alphaNAC), a dynamic component of the ribosomal exit tunnel in eukaryotic cells. Taken together, these findings suggest that alpha-taxilin plays multiple roles in the generation and maintenance of neurons through modulation of the NAC-mediated translational machinary and/or the syntaxin-mediated vesicle traffic in the soma.