Comparative analysis of the priming effect of human interferon-gamma, -alpha, and -beta on synergism with muramyl dipeptide analog for anti-tumor expression of human blood monocytes

Freshly isolated human peripheral blood monocytes from healthy volunteers were not cytotoxic to allogeneic A375 melanoma cells, but they were activated to the cytotoxic state by incubation in vitro with either des-methyl muramyl dipeptide (norMDP; minimal effective dose, 0.5 micrograms/ml) or recombinant human interferon-gamma (rIFN-gamma; minimal effective dose, 1 U/ml). A combination of subthreshold concentrations of these agents (norMDP, 0.5 micrograms/ml; rIFN-gamma, 10 U/ml) also induced significant cytotoxicity, indicating that the effects of norMDP and rIFN-gamma in monocyte activation are synergistic. Natural human IFN-gamma (nIFN-gamma) and norMDP also had similar synergistic effects. Pretreatment of rIFN-gamma with anti-IFN-gamma antibody completely inhibited its synergistic effect with norMDP in monocyte activation. Because pretreatment of rIFN-gamma and norMDP with polymyxin B did not interfere with their effects in monocyte activation, the preparations were not contaminated with lipopolysaccharide. Moreover, because pretreatment of monocyte monolayers with anti-Leu-11b antibody (anti-natural killer (NK) cell antibody) and complement did not interfere with the synergistic effects of norMDP and rIFN-gamma, whereas pretreatment with anti-Leu-M1 antibody (anti-monocyte antibody) caused complete inhibition of their effects, the observed tumor cytotoxicity of monocyte-rich monolayers was probably not due to a small number of adherent NK cells, but to the stimulation of the monocytes. Natural and recombinant IFN-alpha and IFN-beta at concentrations of greater than or equal to 100 U/ml also induced tumoricidal activity of monocytes, but unlike IFN-gamma, their effects were additive with norMDP, and they had less priming effect than IFN-gamma when they were added before norMDP to monocytes. These findings suggest that recombinant human IFN-gamma has much more synergistic potential with norMDP than IFN-alpha or IFN-beta, and this synergism of rIFN-gamma and norMDP for monocyte activation could be of clinical value in treatment of disseminated malignant diseases, because these compounds are readily available at standardized concentrations.     

Reaction of caa3-type terminal cytochrome oxidase from the thermophilic bacterium PS3 with oxygen and carbon monoxide at low temperatures.

Reaction of O2 and CO with a caa3-type terminal cytochrome oxidase (EC 1.9.3.1) from the thermophilic bacterium PS3 grown with high aeration was studied at low temperatures. The CO recombination at the temperature range studied (-50 degrees C to -80 degrees C) followed first-order kinetics with an activation energy of 29.3 kJ/mol (7.0 kcal/mol). In the presence of O2 at -113 degrees C the photolysed reduced form binds O2 to form an 'oxy' intermediate similar to Compound A. At a higher temperature (-97 degrees C) another intermediate, similar to Compound B, is formed as a result of electron transfer from the enzyme to the liganded O2.     

A case of resectable lung adenocarcinoma associated with sarcoidosis

A 71-year-old woman with uveitis was referred to our hospital for further examination of the possible underlying diseases. In roentgenological examination with plain X-ray and CT scan, hilar and mediastinal lymphadenopathy and a mass shadow in the right upper lung field was observed, whereas fibrotic changes were not obvious in both lung fields. Transbronchial lung biopsy with fiberoptic bronchoscope revealed granulomatous interstitial pneumonia. CD4-positive lymphocytes were increased in bronchoalveolar lavage. The patient was diagnosed as having sarcoidosis. Subsequently, right upper lobectomy was performed, and Stage I lung adenocarcinoma was diagnosed. The patient is under follow up without medication and the disease has been stable for two years. A relationship between epithelioid granulomatosis and malignant diseases is discussed and a review of the literature is given. Since it is still controversial as to the incidence of malignant diseases in sarcoidosis patients, it is important to accumulate data on these associations.     

Purification and characterization of malate:quinone oxidoreductase from thermophilic Bacillus sp. PS3

Several bacteria possess membrane-bound dehydrogenases other than cytosolic dehydrogenases in their respiratory chains. In many cases, the membrane-bound malate:quinone oxidoreductases (MQOs) are essential for growth. However, these MQOs are absent in mammalian mitochondria, and therefore may be a potential drug target for pathogenic bacteria. To characterize the kinetic properties of MQOs, we purified MQO from Bacillus sp. PS3, which is a gram-positive and thermophilic bacterium, and cloned the gene encoding MQO based on the obtained partial N-terminus sequence. Purified MQOs showed a molecular mass of ~90 kDa, which was estimated using gel filtration, and it consists of two subunits with a molecular mass of ~50 kDa. Phylogenetic analysis showed a high similarity to the MQO of the Geobacillus group rather than the Bacillus group. Additionally, the purified enzyme was thermostable and it retained menaquinol reduction activity at high temperatures. Although it is difficult to conduct experiments using menaquinol because of its instability, we were able to measure the oxidase activity of cytochrome bd-type quinol oxidase by using menaquinol-1 by coupling this molecule with the menaquinol reduction reaction using purified MQOs.     

Novel methods for nucleotide length control in double-stranded polyinosinic-polycytidylic acid production using uneven length components

ABSTRACT

Polyinosinic-polycytidylic acid (PIC), a double-stranded RNA that induces innate immunity in mammals, is a candidate immunopotentiator for pharmaceuticals. The potency and adverse effects of PIC are strongly correlated with the nucleotide length, and the inability to precisely control the length in PIC production limits its practical use. Length extension during the annealing process is the major factor underlying the lack of control, but tuning the annealing conditions is insufficient to resolve this issue. In this study, we developed a novel method to produce accurate nucleotide length PIC at an industrial scale. The length extension was significantly suppressed by the assembly of multiple short polyinosinic acid molecules with one long polycytidylic acid molecule. A newly developed PIC, uPIC100-400, demonstrated a reproducible length and better storage stability than that of corresponding evenly structured PIC. Human dsRNA receptors exhibited equivalent responsiveness to uPIC100-400 and the evenly structured PIC with the same length.     

Polysomnographic characteristics and predictors of positional obstructive sleep apnea in Japanese elderly

Sleep and Biological Rhythms - Sleep problems and obstructive sleep apnea (OSA) increase with age and disturb life in old age. Positional therapy is one option to treat OSA, but the differences in...     

Low Incidence of Renal Dysfunction among HIV-Infected Patients on a Tenofovir-Based First Line Antiretroviral Treatment Regimen in Myanmar

Background

Since 2004, Médecins Sans Frontières-Switzerland has provided treatment and care for people living with HIV in Dawei, Myanmar. Renal function is routinely monitored in patients on tenofovir (TDF)-based antiretroviral treatment (ART), and this provides an opportunity to measure incidence and risk factors for renal dysfunction.

Methods

We used routinely collected program data on all patients aged ≥15 years starting first-line TDF-based ART between January 2012 and December 2013. Creatinine clearance (CrCl) was assessed at base line and six-monthly, with renal dysfunction defined as CrCl < 50ml/min/1.73m². We calculated incidence of renal dysfunction and used Cox regression analysis to identify associated risk factors.

Results

There were 1391 patients, of whom 1372 had normal renal function at baseline. Of these, 86 (6.3%) developed renal dysfunction during a median time of follow-up 1.14 years with an incidence rate of 5.4 per 100 person-years: 78 had CrCl between 30–50ml/min/1.73m² and were maintained on TDF–based ART, but 5 were changed to another regimen: 4 because of CrCl <30ml/min/1.73m². Risk factors for renal dysfunction included age ≥45 years, diagnosed diabetes, underlying renal disease, underweight and CD4 count <200cells/mm³. There were 19 patients with baseline renal dysfunction and all continued on TDF-based ART: CrCl stayed between 30–49 ml/min/1.73m² in five patients while the remainder regained normal renal function.

Conclusions

In a resource-poor country like Myanmar, the low incidence of renal toxicity in our patient cohort suggests that routine assessment of CrCl may not be needed and could be targeted to high risk groups if resources permit.