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Objective: Rheumatoid arthritis (RA) is a major autoimmune disease that causes synovitis and joint damage. Although clinical trials have been performed using interleukin-10 (IL-10), an antiinflammatory cytokine, as a potential treatment of RA, the therapeutic effects of IL-10 have been limited, potentially due to insufficient residence in lymphoid organs, where antigen recognition primarily occurs. This study was undertaken to engineer an IL-10-serum albumin (SA) fusion protein and evaluate its effects in 2 murine models of RA., Methods: SA-fused IL-10 (SA-IL-10) was recombinantly expressed. Mice with collagen antibody-induced arthritis (n = 4-7 per group) or collagen-induced arthritis (n = 9-15 per group) were injected intravenously with wild-type IL-10 or SA-IL-10, and the retention of SA-IL-10 in the lymph nodes (LNs), immune cell composition in the paws, and therapeutic effect of SA-IL-10 on mice with arthritis were assessed., Results: SA fusion to IL-10 led to enhanced accumulation in the mouse LNs compared with unmodified IL-10. Intravenous SA-IL-10 treatment restored immune cell composition in the paws to a normal status, elevated the frequency of suppressive alternatively activated macrophages, reduced IL-17A levels in the paw-draining LN, and protected joint morphology. Intravenous SA-IL-10 treatment showed similar efficacy as treatment with an anti-tumor necrosis factor antibody. SA-IL-10 was equally effective when administered intravenously, locally, or subcutaneously, which is a benefit for clinical translation of this molecule., Conclusion: SA fusion to IL-10 is a simple but effective engineering strategy for RA therapy and has potential for clinical translation., (© 2020, American College of Rheumatology.)

Background: The novel cap-dependent endonuclease inhibitor baloxavir marboxil was approved in February 2018 for the treatment of influenza virus infection in Japan. In vitro studies have revealed that an I38T substitution in the polymerase acidic subunit (PA) is associated with reduced susceptibility of influenza viruses to baloxavir., Objectives: Development of a rapid and simple method for monitoring influenza A(H1N1)pdm09, A(H3N2), and B viruses possessing the I38T substitution in PA., Methods: Three assays were developed based on RNase H2-dependent PCR (rhPCR) and named A/H1pdm PA_I38T rhPCR, A/H3 PA_I38T rhPCR, and B PA_I38T rhPCR. The assays were evaluated using cDNAs synthesized from in vitro-transcribed PA gene RNA controls, RNAs purified from viruses isolated in the 2017/2018 and 2018/2019 influenza seasons, and RNAs purified from clinical specimens collected in the 2018/2019 influenza season., Results: The assays developed in this study accurately discriminated PA I38 and PA T38 with high sensitivity., Conclusions: Our assays should be considered a powerful tool for monitoring the emergence of baloxavir-resistant influenza viruses., (© 2020 The Authors. Influenza and Other Respiratory Viruses published by John Wiley & Sons Ltd.)

Liver performs several important functions; however, predicting its functions is difficult. Methods of analyzing gene expression profiles, for example, microarray, provide functional information of tissues. Liver and peripheral blood leukocytes (PBLs) were collected from Holstein cows subjected to two different physiological conditions (non-pregnant and pregnant), and PBLs were fractionated by gradient cell separation. RNA from PBLs and liver were applied to oligo-DNA microarray and reverse transcription-quantitative polymerase chain reaction (RT-qPCR). It revealed a group of stable bovine liver genes under constant physiological conditions. When they applied to physiological conditions including non-pregnant and pregnant, the profiles of some genes in liver were consistent with those in PBLs. Microarray data subjected to a principal component analysis (PCA) showed that the hepatic gene expression profiles were more consistent with those of granulocytes than mononuclear cells. The relationship of gene profiles in liver with granulocytes was confirmed by RT-qPCR and hierarchical cluster analysis. Gene profiles of granulocytes were more reliable than those of mononuclear cells, which reflected liver functions. These results suggest that the genes expressed in PBLs, particularly granulocytes, may be convenient bioindicators for the diagnosis of clinical disorder and/or detecting aberration of liver functions in cows subjected to different physiological conditions., (© 2019 Japanese Society of Animal Science.)

We characterized influenza A(H1N1)pdm09 isolates from large-scale outbreaks that occurred in Nepal and India in early 2015. Although no specific viral features, which may have caused the outbreaks, were identified, an S84N substitution in hemagglutinin was frequently observed. Chronological phylogenetic analysis revealed that these Nepalese and Indian viruses possessing the S84N substitution constitute potential ancestors of the novel genetic subclade 6B.1 virus that spread globally in the following (2015/16) influenza season. Thus, active surveillance of circulating influenza viruses in the Southern Asia region, including Nepal and India, would be beneficial for detecting novel variant viruses prior to their worldwide spread., (© 2017 The Authors. Influenza and Other Respiratory Viruses Published by John Wiley & Sons Ltd.)

Background: Adiponectin is an adipocyte-derived hormone that affects regulation of metabolic syndrome such as insulin resistance, type-2 diabetes, and obesity. It functions via seven transmembrane domain receptors [i.e.,adiponectin receptors 1 (AdipoR1) and 2 (AdipoR2)] that have been scarcely investigated in non-human primates., Methods: Molecular cloning of cDNAs for adiponectin, AdipoR1, and AdipoR2 that included the whole protein-coding region in the Japanese monkey, Macaca fuscata, was carried out. Tissue-specific expression of respective genes was analyzed with Northern blot hybridization., Results: The essential Cys36 and four lysine residues in adiponectin, and transmembrane-spanning domains in AdipoR1 and AdipoR2 appear well conserved. While adiponectin mRNA is expressed only in adipose tissues, AdipoR1 mRNA was found to be expressed in various tissues including the brain., Conclusions: These results significantly add to the understanding of the molecular basis of obesity-related adipokines and their receptors in nonhuman primates.

Background: For veterinary management of non-human primates in captivity, and conservation of wild-living primates, management of their health risks is necessary. Incidences of pathogenic bacteria in the fecal specimens are considered as one of the useful indicators for non-invasive health monitoring., Methods: We carried out the detection of Clostridium perfringens in feces from captive and wild chimpanzees by the rapid polymerase chain reaction method., Results: The bacterium was detected in most fecal specimens (80%) in captive chimpanzees. Contrarily, the detection rate in the wild chimpanzees was low, with 23% (n = 12) of 53 fecal samples from the Bossou group, Guinea, and 1.2% (1/81) in the Mahale group, Tanzania., Conclusions: These results show that the intestinal microflora differs between Pan populations under various living conditions, being influenced by their diet and environment.

Background: Japanese monkey, Macaca fuscata, is recognized as the monkey species inhabiting the northernmost area in the world, and thus likely to possess unique fat-depositing mechanisms to resist cold weather in winter. We report that obese females are present in the Wakasa group of Japanese monkey reared in an open enclosure of the Primate Research Institute, Kyoto University., Methods and Results: Eight of 12 females were categorized as obese, showing percentage body fat of over 22%. The levels of serum leptin (mean +/- SD, 4.9 +/- 2.3 ng/ml) measured in these obese monkeys were significantly higher than those of non-obese peers of the same group (n = 4; 1.2 +/- 0.5 ng/ml) and another Japanese monkey group (Takahama, n = 14; 0.8 +/- 0.25 ng/ml); however, serum levels of adiponectin, insulin, glucose, hemoglobin A1c, and fructosamine did not differ between obese and non-obese monkeys. Few serum lipid parameters such as triglyceride and cholesterol showed lower levels in obese monkeys than their non-obese peers., Conclusions: These results show that these obese monkeys in the Wakasa group have not developed obesity-related diseases/disorders such as diabetes. In the Wakasa group, the frequency of obese individuals was high in some maternal lineages, suggesting that genetic factors responsible for obesity may have been inherited in these lineages.

Pepsinogen levels in ape stomachs were comparable to those in macaques and significantly higher than those in the stomachs of other mammals, including carnivores and ruminants. The occurrence of multiple forms of pepsinogens was remarkable. Nine, sixteen, eight, and fourteen pepsinogens were purified or partially purified from the gastric mucosa of a gibbon, orang-utan, gorilla, and chimpanzee, respectively. Most of these were type-A pepsinogens, and only one type-C pepsinogen was identified in each ape. The two types could be readily distinguished by staining for proteolytic activity on polyacrylamide gel electrophoresis (PAGE) in the presence/absence of pepstatin. Type-A pepsinogens were further divided into two subtypes. One subtype, constituting a major group of pepsinogens in apes, exhibited high hemoglobin-digestive activity. The other subtype was specified by a relatively high content of Lys and low hemoglobin-digestive activity. It is likely that pepsinogen-A genes have been duplicated several times as hominoids, including humans, evolved in the primate lineage. The presence of multiple pepsinogens in apes might be advantageous in the efficient digestion of a wide variety of foods.

A cDNA encoding the proform of Bombyx cysteine proteinase (BCP) was expressed at a high level in Escherichia coli using the T7 polymerase expression system. The insoluble recombinant zymogen was solubilized and renatured by modifying a method applied to human pro-cathepsin L. Like the natural BCP precursor, the recombinant proenzyme was spontaneously converted to an active proteinase at pH 3.75. A deletion in the central region of the propeptide resulted in much loss of the activity, suggesting that the propeptide is essential for proper folding during renaturation. In contrast, the renatured mature form of recombinant BCP was not active but regained activity by including the propeptide in the renaturing buffer, suggesting that the propeptide, acting as an intramolecular chaperone, promotes refolding of the associated proteinase domain into an active conformation. The mature form of natural BCP rapidly lost its activity at neutral pH, whereas its proform was stable. The mature enzyme retained some activity in the presence of the propeptide. Arch., (Copyright 1999 Wiley-Liss, Inc.)

Protein inhibitors capable of inhibiting BCP (Bombyx cysteine proteinase) were found in the larval-pupal hemolymph of Bombyx mori. Two forms of the inhibitors, named BCPI (BCP inhibitor) alpha and BCPI beta, were purified from the pupal hemolymph by heat treatment and column chromatographies on CM-cellulose, Toyopearl HW-50, Phenyl-Sepharose, and Mono Q. Purified BCPI beta gave a single protein band with a molecular mass of 10,500 daltons on SDS-PAGE. BCPI alpha is mostly composed of the same molecular mass protein as BCPI beta. Both forms were inhibitory towards other cysteine proteinases such as cathepsins L,B and papain but had no effects on trypsin and pepsin. Both forms inhibited the processing of the enzymatically inactive proform of BCP (pro-BCP) to the activated mature BCP. BCPI alpha and BCPI beta shared many other features such as molecular mass determined by gel filtration, antigenicity, and HPLC profiles. NH(2)-terminal amino acid sequencing of the purified inhibitors revealed that three amino acid residues were different in the BCPI alpha and BCPI beta sequences, all others being identical. The hemolymph BCP inhibitor increased activity approximately four- to fivefold at the time of spinning and maintained this level of activity during pupation., (Copyright 1999 Wiley-Liss, Inc.)

Five pepsinogens were purified from the gastric mucosa of eight species of Asian macaques. The chromatographic behavior of each pepsinogen was essentially the same but differed from human and other mammalian pepsinogens. The major pepsinogen in each species was pepsinogen A-1, accounting for 29-48% of the total. Amino acid compositions and some enzymatic properties of derived pepsins were similar for the various monkey species. This high degree of similarity confirms that these species are closely related to one another.

Two types of nerve cells, sensory and ganglion cells, were identified in the epidermis of the hypostome of Pelmatohydra robusta by light and electron microscopy. In the study of distribution of these cells, the presence of a circumhypostomal nerve ring in the epidermis was revealed, although hydras have been considered to possess only a diffuse nervous system or socalled nerve net. The nerve ring, which encircled the hypostome, was constituted by several clusters of ganglion cells, thick bundles of many neurites connecting these clusters, and a small number of individual ganglion cells located along the bundles. In the nerve ring, some of the lamellae protruding from the ganglion cells were frequently myelinated and wrapped the cell bodies of neighboring ganglion cells, and other lamellae were arranged in concentric circles., (Copyright © 1984 Wiley-Liss, Inc.)