Lavannya Sabharwal

Inflammation Amplifier, a New Paradigm in Cancer Biology

Tumor-associated inflammation can induce various molecules expressed from the tumors themselves or surrounding cells to create a microenvironment that potentially promotes cancer development. Inflammation, particularly chronic inflammation, is often linked to cancer development, even though its evolutionary role should impair nonself objects including tumors. The inflammation amplifier, a hyperinducer of chemokines in nonimmune cells, is the principal machinery for inflammation and is activated by the simultaneous stimulation of NF-κB and STAT3. We have redefined inflammation as local activation of the inflammation amplifier, which causes an accumulation of various immune cells followed by dysregulation of local homeostasis. Genes related to the inflammation amplifier have been genetically associated with various human inflammatory diseases. Here, we describe how cancer-associated genes, including interleukin (IL)-6, Ptgs2, ErbB1, Gas1, Serpine1, cMyc, and Vegf-α, are strongly enriched in genes related to the amplifier. The inflammation amplifier is activated by the stimulation of cytokines, such as TNF-α, IL-17, and IL-6, resulting in the subsequent expression of various target genes for chemokines and tumor-related genes like BCL2L11, CPNE7, FAS, HIF1-α, IL-1RAP, and SOD2. Thus, we conclude that inflammation does indeed associate with the development of cancer. The identified genes associated with the inflammation amplifier may thus make potential therapeutic targets of cancers.

Temporal Expression of Growth Factors Triggered by Epiregulin Regulates Inflammation Development

In this study, we investigated the relationship between several growth factors and inflammation development. Serum concentrations of epiregulin, amphiregulin, betacellulin, TGF-α, fibroblast growth factor 2, placental growth factor (PLGF), and tenascin C were increased in rheumatoid arthritis patients. Furthermore, local blockades of these growth factors suppressed the development of cytokine-induced arthritis in mice by inhibiting chemokine and IL-6 expressions. We found that epiregulin expression was early and followed by the induction of other growth factors at different sites of the joints. The same growth factors then regulated the expression of epiregulin at later time points of the arthritis. These growth factors were increased in patients suffering from multiple sclerosis (MS) and also played a role in the development of an MS model, experimental autoimmune encephalomyelitis. The results suggest that the temporal expression of growth factors is involved in the inflammation development seen in several diseases, including rheumatoid arthritis and MS. Therefore, various growth factor pathways might be good therapeutic targets for various inflammatory diseases.

The Gateway Reflex, which is mediated by the inflammation amplifier, directs pathogenic immune cells into the CNS

The brain-blood barrier (BBB) tightly limits immune cell migration into the central nervous system (CNS), avoiding unwanted inflammation under the normal state. However, immune cells can traverse the BBB when inflammation occurs within the CNS, suggesting a certain signal that creates a gateway that bypasses the BBB might exist. We revealed the inflammation amplifier as a mechanism of this signal, and identified dorsal vessels of the fifth lumber (L5) spinal cord as the gateway. The inflammation amplifier is driven by a simultaneous activation of NF-κB and STATs in non-immune cells, causing the production of a large amount of inflammatory chemokines to open the gateway at L5 vessels. It was found that the activation of the amplifier can be modulated by neural activation and artificially operated by electric pulses followed by establishment of new gateways, Gateway Reflex, at least in mice. Furthermore, genes required for the inflammation amplifier have been identified and are highly associated with various inflammatory diseases and disorders in the CNS. Thus, physical and/or pharmacological manipulation of the inflammation amplifier holds therapeutic value to control neuro-inflammation.

The gateway theory: bridging neural and immune interactions in the CNS.

The central nervous system (CNS) is considered an immune-privileged tissue protected by a specific vessel structure, the blood-brain barrier (BBB). Upon infection or traumatic injury in the CNS, the BBB is breached, and various immune cells are recruited to the affected area. In the case of autoimmune diseases in the CNS like multiple sclerosis (MS), autoreactive T cells against some CNS-specific antigens can theoretically attack neurons throughout the CNS. The affected CNS regions in MS patients can be detected as multiple focal plaques in the cerebrum, thoracic cord, and other regions. Vision problems are often associated with the initial phase of MS, suggesting a disturbance in the optic nerves. These observations raise the possibility that there exist specific signals that direct autoreactive T cells past the BBB and into particular sites of the CNS. Using a mouse model of MS, experimental autoimmune encephalomyelitis (EAE), we recently defined the mechanism of the pathogenesis in which regional neural stimulations modulate the status of the blood vessel endothelium to allow the invasion of autoreactive T cells into specific sites of the CNS via the fifth lumbar cord. This gate for autoreactive T cells can be artificially manipulated by removing gravity forces on the hind legs or by electric pulses to the soleus muscles, quadriceps, and triceps of mice, resulting in an accumulation of autoreactive T cells in the intended regions via the activation of regional neurons. Gating blood vessels by regional neural stimulations, a phenomenon we call the gateway theory, has potential therapeutic value not only in preventing autoimmunity, but also in augmenting the effects of cancer immunotherapies.

Regulation of immune cell infiltration into the CNS by regional neural inputs explained by the gate theory.

The central nervous system (CNS) is an immune-privileged environment protected by the blood-brain barrier (BBB), which consists of specific endothelial cells that are brought together by tight junctions and tight liner sheets formed by pericytes and astrocytic end-feet. Despite the BBB, various immune and tumor cells can infiltrate the CNS parenchyma, as seen in several autoimmune diseases like multiple sclerosis (MS), cancer metastasis, and virus infections. Aside from a mechanical disruption of the BBB like trauma, how and where these cells enter and accumulate in the CNS from the blood is a matter of debate. Recently, using experimental autoimmune encephalomyelitis (EAE), an animal model of MS, we found a “gateway” at the fifth lumber cord where pathogenic autoreactive CD4+ T cells can cross the BBB. Interestingly, this gateway is regulated by regional neural stimulations that can be mechanistically explained by the gate theory. In this review, we also discuss this theory and its potential for treating human diseases.

KDEL receptor 1 regulates T-cell homeostasis via PP1 that is a key phosphatase for ISR

KDEL receptors are responsible for retrotransporting endoplasmic reticulum (ER) chaperones from the Golgi complex to the ER. Here we describe a role for KDEL receptor 1 (KDELR1) that involves the regulation of integrated stress responses (ISR) in T cells. Designing and using an N-ethyl-N-nitrosourea (ENU)-mutant mouse line, T-Red (naïve T-cell reduced), we show that a point mutation in KDELR1 is responsible for the reduction in the number of naïve T cells in this model owing to an increase in ISR. Mechanistic analysis shows that KDELR1 directly regulates protein phosphatase 1 (PP1), a key phosphatase for ISR in naïve T cells. T-Red KDELR1 does not associate with PP1, resulting in reduced phosphatase activity against eIF2α and subsequent expression of stress responsive genes including the proapoptotic factor Bim. These results demonstrate that KDELR1 regulates naïve T-cell homeostasis by controlling ISR.

Breakpoint Cluster Region–Mediated Inflammation Is Dependent on Casein Kinase II

The breakpoint cluster region (BCR) is known as a kinase and cause of leukemia upon fusing to Abl kinase. In this study, we demonstrate that BCR associated with the α subunit of casein kinase II (CK2α), rather than BCR itself, is required for inflammation development. We found that BCR knockdown inhibited NF-κB activation in vitro and in vivo. Computer simulation, however, suggested that the putative BCR kinase domain has an unstable structure with minimal enzymatic activity. Liquid chromatography–tandem mass spectrometry analysis showed that CK2α associated with BCR. We found the BCR functions are mediated by CK2α. Indeed, CK2α associated with adaptor molecules of TNF-αR and phosphorylated BCR at Y177 to establish a p65 binding site after TNF-α stimulation. Notably, p65 S529 phosphorylation by CK2α creates a p300 binding site and increased p65-mediated transcription followed by inflammation development in vivo. These results suggest that BCR-mediated inflammation is dependent on CK2α, and the BCR–CK2α complex could be a novel therapeutic target for various inflammatory diseases.

Strong TCR-mediated signals suppress integrated stress responses induced by KDELR1 deficiency in naive T cells

KDEL receptor 1 (KDELR1) regulates integrated stress responses (ISR) to promote naive T-cell survival in vivo. In a mouse line having nonfunctional KDELR1, T-Red (naive T-cell reduced) mice, polyclonal naive T cells show excessive ISR and eventually undergo apoptosis. However, breeding T-Red mice with TCR-transgenic mice bearing relatively high TCR affinity rescued the T-Red phenotype, implying a link between ISR-induced apoptosis and TCR-mediated signaling. Here, we showed that strong TCR stimulation reduces ISR in naive T cells. In mice lacking functional KDELR1, surviving naive T cells expressed significantly higher levels of CD5, a surrogate marker of TCR self-reactivity. In addition, higher TCR affinity/avidity was confirmed using a tetramer dissociation assay on the surviving naive T cells, suggesting that among the naive T-cell repertoire, those that receive relatively stronger TCR-mediated signals via self-antigens survive enhanced ISR. Consistent with this observation, weak TCR stimulation with altered peptide ligands decreased the survival and proliferation of naive T cells, whereas stimulation with ligands having higher affinity had no such effect. These results suggest a novel role of TCR-mediated signals in the attenuation of ISR in vivo.

Role of Cytokine-Mediated Crosstalk between T Cells and Nonimmune Cells in the Pathophysiology of Multiple Sclerosis

Abstract Multiple sclerosis (MS) is a progressive demyelinating disease associated with chronic inflammation in the central nervous system (CNS). Genetic linkage is found in genes related to T cell function, and T cell infiltration is evident in MS lesions, suggesting a pathophysiological role for these cells. In the MS animal model experimental autoimmune encephalomyelitis (EAE), activated CD4+ and CD8+ T cells that recognize CNS antigens can induce MS-like symptoms. Various cytokines from these T cells trigger inflammation, critically contributing to the pathogenesis of the CNS disease. Evidence suggests an important role for cytokine-mediated crosstalk between T cells and nonimmune cells, including endothelial cells, glial cells, and fibroblasts in enhancing CNS inflammation. Counterbalancing T cell-induced inflammation, certain regulatory CD4+ and CD8+ T cell subsets can suppress CNS disease progression. Here, we discuss the pathophysiological role of T cells during MS and EAE, including neuro–immune interactions that allow T cells to invade the CNS.

Presenilin 1 Regulates NF-κB Activation via Association with Breakpoint Cluster Region and Casein Kinase II

We recently reported that NF-κB–mediated inflammation caused by breakpoint cluster region (BCR) is dependent on the α subunit of casein kinase II (CK2α) complex. In the current study, we demonstrate that presenilin 1 (Psen1), which is a catalytic component of the γ-secretase complex and the mutations of which are known to cause familial Alzheimer disease, acts as a scaffold of the BCR–CK2α–p65 complex to induce NF-κB activation. Indeed, Psen1 deficiency in mouse endothelial cells showed a significant reduction of NF-κB p65 recruitment to target gene promoters. Conversely, Psen1 overexpression enhanced reporter activation under NF-κB responsive elements and IL-6 promoter. Furthermore, the transcription of NF-κB target genes was not inhibited by a γ-secretase inhibitor, suggesting that Psen1 regulates NF-κB activation in a manner independent of γ-secretase activity. Mechanistically, Psen1 associated with the BCR–CK2α complex, which is required for phosphorylation of p65 at serine 529. Consistently, TNF-α–induced phosphorylation of p65 at serine 529 was significantly decreased in Psen1-deficient cells. The association of the BCR–CK2α–p65 complex was perturbed in the absence of Psen1. These results suggest that Psen1 functions as a scaffold of the BCR–CK2α–p65 complex and that this signaling cascade could be a novel therapeutic target for various chronic inflammation conditions, including those in Alzheimer disease.