IFN-γ, a proinflammatory cytokine produced primarily by T cells and NK cells, activates macrophages and engages mechanisms to control pathogens. Although there is evidence of IFN-γ production by murine macrophages, IFN-γ production by normal human macrophages and their subsets remains unknown. Herein, we show that human M1 macrophages generated by IFN-γ and IL-12– and IL-18–stimulated monocyte-derived macrophages (M0) produce significant levels of IFN-γ. Further stimulation of IL-12/IL-18–primed macrophages or M1 macrophages with agonists for TLR-2, TLR-3, or TLR-4 significantly enhanced IFN-γ production in contrast to the similarly stimulated M0, M2a, M2b, and M2c macrophages. Similarly, M1 macrophages generated from COVID-19–infected patients’ macrophages produced IFN-γ that was enhanced following LPS stimulation. The inhibition of M1 differentiation by Jak inhibitors reversed LPS-induced IFN-γ production, suggesting that differentiation with IFN-γ plays a key role in IFN-γ induction. We subsequently investigated the signaling pathway(s) responsible for TLR-4–induced IFN-γ production in M1 macrophages. Our results show that TLR-4–induced IFN-γ production is regulated by the ribosomal protein S6 kinase (p70S6K) through the activation of PI3K, the mammalian target of rapamycin complex 1/2 (mTORC1/2), and the JNK MAPK pathways. These results suggest that M1-derived IFN-γ may play a key role in inflammation that may be augmented following bacterial/viral infections. Moreover, blocking the mTORC1/2, PI3K, and JNK MAPKs in macrophages may be of potential translational significance in preventing macrophage-mediated inflammatory diseases.

Small interfering RNA (siRNA) is a critical loss-of-function tool for elucidating the role of genes in biomedical studies. The effective use of siRNA needs transfection technology that delivers siRNA into the correct location of target cells, especially those which are extremely difficult to transfect. Macrophages, which play an important role in the pathogenesis of many diseases, are known to be extremely hard to transfect. Thus, to elucidate the functions of genes in human macrophage biology, it is essential to devise technology for efficient siRNA transfection. However, a fast and efficient method for siRNA transfection in primary human macrophages has not been reported. The siRNA transfection is a tug-of-war between transfection rate and cytotoxicity. A higher transfection rate is generally accompanied with increased cytotoxicity, therefore, choosing a transfection reagent that limits cell death while maintain a desirable transfection rate is important. In this study, we employed auto-analysis function of the IncuCyte to devise a fast and cost-saving technology for efficient transfection of adherent cells and particularly human macrophages. We show that DharmaFECT3 transfection reagent from Dharmacon was the most efficient in transfecting primary human monocyte-derived macrophages and PMA-differentiated U937 cells, whereas other transfection reagents tested were cytotoxic. This method exhibited approximately 85% transfection efficiency in human macrophages and U937 cells. Moreover, siRNA silencing of Bax with this technique effectively protected primary human macrophages and PMA-differentiated U937 cells against Resveratrol-induced cell death. In addition, this method inherently takes the balance between transfection rate and cytotoxicity of siRNA transfection reagents into consideration.

In this article, we compiled 13 etiological theories, 15 characteristics, and 81 observations/phenomena of peptic ulcers, reported in reproducible, peer-reviewed studies from the literature, to reflect the historical evolution of studies on peptic ulcers and to provide a multidisciplinary view of this disease. This data was collected during the systematic review of topics on peptic ulcers including genetics, etiology, epidemiology, psychology, anatomy, neurology, bacteriology, pathology, and clinical statistics. The data curated herein was extracted via application of recently published basic theories and methodologies.

Inflammatory macrophages have been implicated in many diseases, including rheumatoid arthritis and inflammatory bowel disease. Therefore, targeting macrophage function and activation may represent a potential strategy to treat macrophage-associated diseases. We have previously shown that IFN-γ–induced differentiation of human M0 macrophages toward proinflammatory M1 state rendered them highly susceptible to the cytocidal effects of second mitochondria-derived activator of caspases mimetics (SMs), antagonist of the inhibitors of apoptosis proteins (IAPs), whereas M0 and anti-inflammatory M2c macrophages were resistant. In this study, we investigated the mechanism governing SM-induced cell death during differentiation into M1 macrophages and in polarized M1 macrophages. IFN-γ stimulation conferred on M0 macrophages the sensitivity to SM-induced cell death through the Jak/STAT, IFN regulatory factor-1, and mammalian target of rapamycin complex-1 (mTORC-1)/ribosomal protein S6 kinase pathways. Interestingly, mTORC-1 regulated SM-induced cell death independent of M1 differentiation. In contrast, SM-induced cell death in polarized M1 macrophages is regulated by the mTORC-2 pathway. Moreover, SM-induced cell death is regulated by cellular IAP (cIAP)-2, receptor-interacting protein kinase (RIPK)-1, and RIPK-3 degradation through mTORC activation during differentiation into M1 macrophages and in polarized M1 macrophages. In contrast to cancer cell lines, SM-induced cell death in M1 macrophages is independent of endogenously produced TNF-α, as well as the NF-κB pathway. Collectively, selective induction of cell death in human M1 macrophages by SMs may be mediated by cIAP-2, RIPK-1, and RIPK-3 degradation through mTORC activation. Moreover, blocking cIAP-1/2, mTORC, or IFN regulatory factor-1 may represent a promising therapeutic strategy to control M1-associated diseases.

Peptic ulcers, including duodenal and gastric ulcers, are currently studied as an infectious disease caused by Helicobacter pylori. However, this etiology cannot elucidate the major characteristics and observations/phenomena of the disease, as well as the roles of gastric acid and NSAIDs. Additionally, many patients with severe ulcer symptoms have no infection, suggesting bacterial eradication cannot prevent relapses in all ulcer patients. To address these challenges, a recently published Complex Causal Relationship with its accompanying methodologies was applied to analyze the existing data. Peptic ulcers were identified as a psychosomatic disease triggered by psychological stress, where Helicobacter pylori plays a secondary role in only the late phase of ulceration. An integration of five major etiological theories in history illustrated the entire pathogenesis of peptic ulcers, which addressed all the characteristics, observations/phenomena, controversies, and mysteries of the disease in a series of 6 articles. This first article focuses on the pathogenesis of duodenal ulcers. The hyperplasia and hypertrophy of gastrin and parietal cells induced by chronic stress potentiate the individual’s response to immediate stress, resulting in the hypersecretion of gastric acid and eventually, duodenal ulceration. This psychopathological mechanism resolved all the controversies associated with Helicobacter pylori and explicitly elucidated 7 characteristics of duodenal ulcers, as well as 42 observations/phenomena. The roles of gastric acid, Helicobacter pylori, and NSAIDs in duodenal ulcers were also identified. The effectiveness validated the etiology of duodenal ulcers identified by the Complex Causal Relationship, indicating stress management is essential for a complete cure of the disease without relapse.