Oral 5-aminolevulinic acid combined with sodium ferrous citrate prevents blood-aqueous barrier breakdown after anterior chamber paracentesis in healthy beagle dogs.

This study investigated the preventive effect of 5-aminolevulinic acid combined with sodium ferrous citrate (5-ALA/SFC) on blood-aqueous barrier (BAB) breakdown induced after anterior chamber paracentesis (ACP) in beagles. 5-ALA/SFC (1/0.64 mg/kg or 3/1.92 mg/kg) or carprofen (4.0 mg/kg) was orally administered daily for 7 days prior to ACP. Then, a sample of the aqueous humor (AH) was collected from one eye via ACP (first sample) and again 60 min later (second sample). The protein and prostaglandin E2 (PGE2) concentrations in both samples were measured. Compared with the control group, high-dose 5-ALA/SFC and carprofen significantly reduced the AH protein and PGE2 concentrations in the second sample. Our findings suggest that 5-ALA/SFC suppresses BAB breakdown in dogs.

The effect of 5-aminolevulinic acid on canine peripheral blood mononuclear cells.

5-aminolevulinic acid (ALA) is a natural amino acid and a product of the first heme synthesis pathway in mitochondria. Its immunomodulatory effects have garnered recent attention for their potential application to cancer, inflammation, and autoimmune diseases in humans. A supplement containing ALA is now available in Japan to enhance ATP synthesis via mitochondrial activity. However, how ALA affects canine immunity is unclear. Here we studied the effects of ALA on peripheral blood mononuclear cells (PBMCs) from healthy dogs in vitro. Heme oxygenase-1 (HO-1) protein was expressed in Madin-Darby canine kidney (MDCK) cells and PBMCs treated with ALA and ferrous sodium citrate (SFC), which showed that ALA works in dogs as well as humans. ALA also induced concanavalin A (ConA)-stimulated PBMCs to produce significantly more interferon-gamma (IFN-γ). Next-generation RNA sequencing (RNA-seq) revealed that ALA enhanced T cell immunity among Th1, Th2, and Th17 subsets, especially the IL-17 signaling pathway. We then confirmed that ALA promoted interleukin (IL)- 17A production in ConA-stimulated PBMCs. Together, these findings indicate that ALA promotes heme synthesis in mitochondria and enhances ConA-induced T cell immune responses in canine PBMCs.

High-throughput laboratory evolution reveals evolutionary constraints in Escherichia coli.

Understanding the constraints that shape the evolution of antibiotic resistance is critical for predicting and controlling drug resistance. Despite its importance, however, a systematic investigation of evolutionary constraints is lacking. Here, we perform a high-throughput laboratory evolution of Escherichia coli under the addition of 95 antibacterial chemicals and quantified the transcriptome, resistance, and genomic profiles for the evolved strains. Utilizing machine learning techniques, we analyze the phenotype-genotype data and identified low dimensional phenotypic states among the evolved strains. Further analysis reveals the underlying biological processes responsible for these distinct states, leading to the identification of trade-off relationships associated with drug resistance. We also report a decelerated evolution of ß-lactam resistance, a phenomenon experienced by certain strains under various stresses resulting in higher acquired resistance to ß-lactams compared to strains directly selected by ß-lactams. These findings bridge the genotypic, gene expression, and drug resistance gap, while contributing to a better understanding of evolutionary constraints for antibiotic resistance.

Improvement Effect of 5-Aminolevulinic Acid on Hyperlipidemia in Miniature Schnauzer Dogs: An Open Study in 5 Cases of One Pedigree.

This is the first study to examine the long-term effect of 5-aminolevulinic acid mainly on serum lipoproteins in dogs with hyperlipidemia. We studied 5 Miniature Schnauzer cases whose fasting serum total triglyceride and very-low-density lipoprotein of triglyceride levels were extremely high (635 ± 116 and 520 ± 92 mg/dL, respectively). Although the total cholesterol values were normal, the very-low-density lipoprotein of cholesterol level was high (49 ± 7 mg/dL). Each dog received a 5-aminolevulinic acid supplement (5 mg/day) orally for 6 months. The mean values of total triglyceride, very-low-density lipoprotein of both triglyceride and cholesterol decreased significantly after the treatment period (319 ± 29, 245 ± 18, and 27 ± 2 mg/dL, respectively, P < 0.05). Our present results may present evidence that 5-ALA administration contributes to improvement of hyperlipidemia in Miniature Schnauzer.

High-throughput identification of the sensitivities of an Escherichia coli ΔrecA mutant strain to various chemical compounds.

Antibiotic resistance is considered a global threat to public health. Adaptive resistance mutations and the acquisition of resistance genes by horizontal gene transfer are known to be facilitated by the RecA-dependent SOS response during antibiotic treatment, making RecA inhibitors promising agents for the prevention of antibiotic resistance. However, the impact of RecA inactivation on antibiotic sensitivities remains unclear. Therefore, in this study, we performed high-throughput screening to determine the minimum inhibitory concentrations (MICs) of 217 chemicals, including both antibiotics and toxic chemicals of unknown drug action, in the wild-type MDS42 and the ΔrecA mutant strains of Escherichia coli. The ΔrecA mutant showed increased sensitivity to DNA-damaging agents, DNA replication inhibitors, and chromate stress, as well as to other chemicals, such as S-(2-aminoethyl)-L-cysteine, L-histidine, ruthenium red, D-penicillamine, carbonyl cyanide 3-chlorophenylhydrazone (CCCP), cerulenin, and L-cysteine. Microarray analysis showed further that the ΔrecA mutant had lower expressions of glnK, nac, and glnLG, which encode nitrogen assimilation regulators, as well as amtB, which encodes an ammonium transporter, compared with the wild type. These findings suggest that the ΔrecA mutation affects not only the SOS response but also amino acid metabolism.

Improvement of isopropanol tolerance of Escherichia coli using adaptive laboratory evolution and omics technologies.

Isopropanol (IPA) is the secondary alcohol that can be dehydrated to yield propylene. To produce IPA using microorganisms, a significant issue is that the toxicity of IPA causes retardation or inhibition of cell growth, decreasing the yield. One possible strategy to overcome this problem is to improve IPA tolerance of production organisms. For the understanding of tolerance to IPA, we performed parallel adaptive laboratory evolution (ALE) of Escherichia coli under IPA stress. To identify the genotypic change during ALE, we performed genome re-sequencing analyses of obtained tolerant strains. To verify which mutations were contributed to IPA tolerance, we constructed the mutant strains and quantify the IPA tolerance of the constructed mutants. From these analyses, we found that five mutations (relA, marC, proQ, yfgO, and rraA) provided the increase of IPA tolerance. To understand the phenotypic change during ALE, we performed transcriptome analysis of tolerant strains. From transcriptome analysis, we found that expression levels of genes related to biosynthetic pathways of amino acids, iron ion homeostasis, and energy metabolisms were changed in the tolerant strains. Results from these experiments provide fundamental bases for designing IPA tolerant strains for industrial purposes.

Changes of epidermal Langerhans cells in skin treated with trichloroacetic acid.

The changes in Langerhans cells (LCs) from normal human skin peeled with 40 and 60% trichloroacetic acid (TCA) or liquid nitrogen , which is known in cryosurgery as a control, were examined using monoclonal antibodies against the CD1a, HLA-DR and Lag in order to examine the immune surveillance system. In the 40% TCA group, the number of CD1a-positive cells decreased gradually until day 7, whereas both HLA-DR- and Lag-positive cells decreased for 12 hours, increased until day 2 and decreased thereafter. In the 60% TCA group, the number of CD1a-, HLA-DR- and Lag-positive cells decreased gradually until day1, increased temporarily until day 2, and decreased again until day 7. There were no significant differences in the decrease of the LCs between the 40% and 60% TCA groups. In both cases the number of LCs on day 7 was statistically lower than before treatment. In the liquid nitrogen group, which served as a control, the LCs decreased gradually and slightly until day 2, and then increased. Taken together, the number of epidermal LCs from TCA-treated skin was reduced more significantly than LCs from liquid nitrogen-treated skin, suggesting a temporary impairment of the skin defense system. Therefore, long-term and frequent TCA peeling will require special attention for potential carcinogenesis.