Inoculation of Chlamydia pneumoniae or Chlamydia trachomatis with ligands that inhibit attachment to host cells reduces infectivity in the mouse model of lung infection: implication for anti-adhesive therapy

Previous studies have shown that the chlamydial glycan contains a high-mannose oligosaccharide, which mediates attachment and infectivity of the organism. Removal of the glycan decreases infectivity in vitro and in vivo. The present study demonstrates that simultaneous inoculation of chlamydial organisms and a ligand that prevents glycan binding reduces lung burden in infected animals.     

Protective effect of pioglitazone on morphine-induced neuroinflammation in the rat lumbar spinal cord

Background

Morphine-induced tolerance is associated with the spinal neuroinflammation. The aim of this study was to explore the effects of oral administration of the pioglitazone, the peroxisome proliferator activated receptor gamma (PPAR-γ) agonist, on the morphine-induced neuroinflammation in the lumbar region of the male Wistar rat spinal cord.

Results

Co-administration of the pioglitazone with morphine not only attenuated morphine-induced tolerance, but also prevented the up-regulation of pro-inflammatory cytokines (tumor necrosis factor alpha, interleukin-1beta, and interleukin 6) and nuclear factor-kappa B activity. Administration of the GW-9662 antagonized the above mentioned effects of the pioglitazone.

Conclusions

It is concluded that oral administration of the pioglitazone attenuates morphine-induced tolerance and the neuroinflammation in the lumbar region of the rat spinal cord. This action of the pioglitazone may be, at least in part, due to an interaction with the spinal pro-inflammatory cytokine expression and the nuclear factor-kappa B activity.     

Disruption of Paneth and goblet cell homeostasis and increased endoplasmic reticulum stress in Agr2−/− mice

Anterior Gradient 2 (AGR2) is a protein disulfide isomerase that plays important roles in diverse processes in multiple cell lineages as a developmental regulator, survival factor and susceptibility gene for inflammatory bowel disease. Here, we show using germline and inducible Agr2−/− mice that Agr2 plays important roles in intestinal homeostasis. Agr2−/− intestine has decreased goblet cell Mucin 2, dramatic expansion of the Paneth cell compartment, abnormal Paneth cell localization, elevated endoplasmic reticulum (ER) stress, severe terminal ileitis and colitis. Cell culture experiments show that Agr2 expression is induced by ER stress, and that siRNA knockdown of Agr2 increases ER stress response. These studies implicate Agr2 in intestinal homeostasis and ER stress and suggest a role in the etiology of inflammatory bowel disease.     

A C-terminal domain in KCC2 confers constitutive K+-Cl- cotransport

The neuron-specific K(+)-Cl(-) cotransporter KCC2 plays a crucial role in determining intracellular chloride activity and thus the neuronal response to gamma-aminobutyric acid and glycine. Of the four KCCs, KCC2 is unique in mediating constitutive K(+)-Cl(-) cotransport under isotonic conditions; the other three KCCs are exclusively swelling-activated, with no isotonic activity. We have utilized a series of chimeric cDNAs to localize the determinant of isotonic transport in KCC2. Two generations of chimeric KCC4-KCC2 cDNAs initially localized this characteristic to within a KCC2-specific expansion of the cytoplasmic C terminus, between residues 929 and 1043. This region of KCC2 is rich in prolines, serines, and charged residues and encompasses two predicted PEST sequences. Substitution of this region in KCC2 with the equivalent sequence of KCC4 resulted in a chimeric KCC that was devoid of isotonic activity, with intact swelling-activated transport. A third generation of chimeras demonstrated that a domain just distal to the PEST sequences confers isotonic transport on KCC4. Mutagenesis of this region revealed that residues 1021-1035 of KCC2 are sufficient for isotonic transport. Swelling-activated K(+)-Cl(-) cotransport is abrogated by calyculin A, whereas isotonic transport mediated by KCC chimeras and KCC2 is completely resistant to this serine-threonine phosphatase inhibitor. In summary, a 15-residue C-terminal domain in KCC2 is both necessary and sufficient for constitutive K(+)-Cl(-) cotransport under isotonic conditions. Furthermore, unlike swelling-activated transport, constitutive K(+)-Cl(-) cotransport mediated by KCC2 is completely independent of serine-threonine phosphatase activity, suggesting that these two modes of transport are activated by distinct mechanisms.     

Type 2 IDI performs better than type 1 for improving lycopene production in metabolically engineered E. coli strains

In this study a comparison was made between type 1 and type 2 isopentenyl diphosphate isomerases (IDI) in improving lycopene production in Escherichia coli. The corresponding genes of Bacillus licheniformis and the host (i _Bl and i _Ec , respectively) were expressed in lycopene producing E. coli strains by pTlyci_Bl and pTlyci_Ec plasmids, under the control of tac promoter. The results showed that the overexpression of i _Ec improved the lycopene production from 33 ± 1 in E. coli Tlyc to 68 ± 3 mg/gDCW in E. coli Tlyci_Ec. In contrast, the expression of i _Bl increased the lycopene production more efficiently up to 80 ± 9 mg/gDCW in E. coli Tlyci_Bl. The introduction of a heterologous mevalonate pathway to elevate the IPP abundance resulted in a lycopene production up to 132 ± 5 mg/gDCW with i _Ec in E. coli Tlyci_Ec-mev and 181 ± 9 mg/gDCW with i _Bl in E. coli Tlyci_Bl-mev, that is, 4 and 5.6 times respectively. When fructose, mannose, arabinose, and acetate were each used as an auxiliary substrate with glycerol, lycopene production was inhibited by different extents. Among auxiliary substrates tested, only citrate was an improving one for lycopene production in all strains with a maximum of 198 ± 3 mg/gDCW in E. coli Tlyci_Bl-mev. It may be concluded that the type 2 IDI performs better than the type 1 in metabolic engineering attempts for isoprenoid production in E. coli. In addition, the metabolic engineering of citrate pathway seems a promising approach to have more isoprenoid accumulation in E. coli.     

Isolation,Culture and Long-Term Maintenance of Primary Mesencephalic Dopaminergic Neurons From Embryonic Rodent Brains

Degeneration of mesencephalic dopaminergic (mesDA) neurons is the pathological hallmark of Parkinson’s diseae. Study of the biological processes involved in physiological functions and vulnerability and death of these neurons is imparative to understanding the underlying causes and unraveling the cure for this common neurodegenerative disorder. Primary cultures of mesDA neurons provide a tool for investigation of the molecular, biochemical and electrophysiological properties, in order to understand the development, long-term survival and degeneration of these neurons during the course of disease. Here we present a detailed method for the isolation, culturing and maintenance of midbrain dopaminergic neurons from E12.5 mouse (or E14.5 rat) embryos. Optimized cell culture conditions in this protocol result in presence of axonal and dendritic projections, synaptic connections and other neuronal morphological properties, which make the cultures suitable for study of the physiological, cell biological and molecular characteristics of this neuronal population.     

Physiological roles of the mitochondrial permeability transition pore

Neurons experience high metabolic demand during such processes as synaptic vesicle recycling, membrane potential maintenance and Ca²⁺ exchange/extrusion. The energy needs of these events are met in large part by mitochondrial production of ATP through the process of oxidative phosphorylation. The job of ATP production by the mitochondria is performed by the F₁F_O ATP synthase, a multi-protein enzyme that contains a membrane-inserted portion, an extra-membranous enzymatic portion and an extensive regulatory complex. Although required for ATP production by mitochondria, recent findings have confirmed that the membrane-confined portion of the c-subunit of the ATP synthase also houses a large conductance uncoupling channel, the mitochondrial permeability transition pore (mPTP), the persistent opening of which produces osmotic dysregulation of the inner mitochondrial membrane, uncoupling of oxidative phosphorylation and cell death. Recent advances in understanding the molecular components of mPTP and its regulatory mechanisms have determined that decreased uncoupling occurs in states of enhanced mitochondrial efficiency; relative closure of mPTP therefore contributes to cellular functions as diverse as cardiac development and synaptic efficacy.     

Isolation, structural characterization and antioxidant activity of a new water-soluble polysaccharide from Acanthophyllum bracteatum roots

ABPS-1, a new water-soluble polysaccharide with molecular weight of 26 kDa and a specific optical rotation of +170° (c 1.0, H₂O), was extracted from the roots of Acanthophyllum bracteatum by warm water and further successively purified through DEAE-cellulose A52 and Sephadex G-100 columns. Monosaccharide analysis revealed that the ABPS-1 was composed of Glc, Gal and Ara with a relative molar ratio of 1.4:5.2:1.0. Its structural features were elucidated by a combination of FT-IR, methylation and GC-MS analysis, periodate oxidation and Smith degradation, partial acid hydrolysis and ¹³C and ¹H NMR spectroscopy. The data obtained indicate that ABPS-1 possessed a backbone of α-(1 → 6)-linked Gal with branches attached to O-2 by α-1 → linked Glc and at O-3 by α-1 → linked Gal and by α-(1 → 3)-linked Ara. The in vitro antioxidant activity showed that ABPS-1 possesses DPPH radical-scavenging activity in a concentration-dependent manner with an EC₅₀ value of 2.6 mg/ml.