Splenic marginal zone B cells restrict Mycobacterium tuberculosis infection by shaping the cytokine pattern and cell-mediated immunity.

Understanding the role of B cells in tuberculosis (TB) is crucial for developing new TB vaccines. However, the changes in B cell immune landscapes during TB and their functional implications remain incompletely explored. Using high-dimensional flow cytometry to map the immune landscape in response to Mycobacterium tuberculosis (Mtb) infection, our results show an accumulation of marginal zone B (MZB) cells and other unconventional B cell subsets in the lungs and spleen, shaping an unconventional B cell landscape. These MZB cells exhibit activated and memory-like phenotypes, distinguishing their functional profiles from those of conventional B cells. Notably, functional studies show that MZB cells produce multiple cytokines and contribute to systemic protection against TB by shaping cytokine patterns and cell-mediated immunity. These changes in the immune landscape are reversible upon successful TB chemotherapy. Our study suggests that, beyond antibody production, targeting the regulatory function of B cells may be a valuable strategy for TB vaccine development.

A Novel Tool to Identify Bactericidal Compounds against Vulnerable Targets in Drug-Tolerant M. tuberculosis found in Caseum.

Caseous necrosis is a hallmark of tuberculosis (TB) pathology and creates a niche for drug-tolerant persisters within the host. Cavitary TB and high bacterial burden in caseum require longer treatment duration. An in vitro model that recapitulates the major features of Mycobacterium tuberculosis (Mtb) in caseum would accelerate the identification of compounds with treatment-shortening potential. We have developed a caseum surrogate model consisting of lysed and denatured foamy macrophages. Upon inoculation of Mtb from replicating cultures, the pathogen adapts to the lipid-rich matrix and gradually adopts a nonreplicating state. We determined that the lipid composition of ex vivo caseum and the surrogate matrix are similar. We also observed that Mtb in caseum surrogate accumulates intracellular lipophilic inclusions (ILI), a distinctive characteristic of quiescent and drug-tolerant Mtb. Expression profiling of a representative gene subset revealed common signatures between the models. Comparison of Mtb drug susceptibility in caseum and caseum surrogate revealed that both populations are similarly tolerant to a panel of TB drugs. By screening drug candidates in the surrogate model, we determined that the bedaquiline analogs TBAJ876 and TBAJ587, currently in clinical development, exhibit superior bactericidal against caseum-resident Mtb, both alone and as substitutions for bedaquiline in the bedaquiline-pretomanid-linezolid regimen approved for the treatment of multidrug-resistant TB. In summary, we have developed a physiologically relevant nonreplicating persistence model that reflects the distinct metabolic and drug-tolerant state of Mtb in caseum. IMPORTANCE M. tuberculosis (Mtb) within the caseous core of necrotic granulomas and cavities is extremely drug tolerant and presents a significant hurdle to treatment success and relapse prevention. Many in vitro models of nonreplicating persistence have been developed to characterize the physiologic and metabolic adaptations of Mtb and identify compounds active against this treatment-recalcitrant population. However, there is little consensus on their relevance to in vivo infection. Using lipid-laden macrophage lysates, we have designed and validated a surrogate matrix that closely mimics caseum and in which Mtb develops a phenotype similar to that of nonreplicating bacilli in vivo. The assay is well suited to screen for bactericidal compounds against caseum-resident Mtb in a medium-throughput format, allowing for reduced reliance on resource intensive animal models that present large necrotic lesions and cavities. Importantly, this approach will aid the identification of vulnerable targets in caseum Mtb and can accelerate the development of novel TB drugs with treatment-shortening potential.

Combination of radiation and interleukin 12 eradicates large orthotopic hepatocellular carcinoma through immunomodulation of tumor microenvironment.

Immunotherapies have shown promising results in certain cancer patients. For hepatocellular carcinoma (HCC), the multiplicity of an immunotolerant microenvironment within both the tumor, and the liver per se may limit the efficacy of cancer immunotherapies. Since radiation induces immunogenic cell death and inflammatory reactions within the tumor microenvironment, we hypothesized that a combination therapy of radiation and lasting local immunostimulating agents, achieved by intratumoral injection of an adenoviral vector encoding interleukin 12, may reverse the immunotolerant microenvironment within a well-established orthotopic HCC toward a state favorable for inducing antitumor immunities. Our data showed that radiation and IL-12 combination therapy (RT/IL-12) led to dramatic tumor regression in animals bearing large subcutaneous or orthotopic HCC, induced systemic effect against distant tumor, and significantly prolonged survival. Radiation monotherapy induced tumor regression at early times but afterwards most tumors regained exponential growth, while IL-12 monotherapy only delayed tumor growth. Mechanistic studies revealed that RT/IL-12 increased expression of MHC class II and co-stimulatory molecules CD40 and CD86 on tumor-infiltrating dendritic cells, suggesting an improvement of their antigen presentation activity. RT/IL-12 also significantly reduced accumulation of tumor-infiltrating myeloid-derived suppressor cells (MDSCs) and impaired their suppressive functions by reducing production of reactive oxygen species. Accordingly, tumor-infiltrating CD8+ T cells and NK cells were significantly activated toward the antitumor phenotype, as revealed by increased expression of CD107a and TNF-α. Together, our data showed that RT/IL-12 treatment could reset the intratumoral immunotolerant state and stimulate activation of antitumor cellular immunity that is capable of eliminating large established HCC tumors.

TREM-1 regulates macrophage polarization in ureteral obstruction.

Chronic kidney disease (CKD) is an emerging worldwide public health problem. Inflammatory cell infiltration and activation during the early stages in injured kidneys is a common pathologic feature of CKD. Here, we determined whether an important inflammatory regulator, triggering receptor expressed on myeloid cells (TREM)-1, is upregulated in renal tissues collected from mouse ureteral obstruction-induced nephritis. TREM-1 is crucial for modulating macrophage polarization, and has a pivotal role in mediating tubular injury and interstitial collagen deposition in obstructive nephritis. Lysates from nephritic kidneys triggered a TREM-1-dependent M1 polarization ex vivo, consistent with the observation that granulocyte-macrophage colony-stimulating factor (GM-CSF)-derived M1 macrophages express higher levels of TREM-1 in comparison with M-CSF-derived cells. Moreover, agonistic TREM-1 cross-link significantly strengthens the inductions of iNOS and GM-CSF in M1 cells. These observations are validated by a strong clinical correlation between infiltrating TREM-1-expressing/iNOS-positive macrophages and renal injury in human obstructive nephropathy. Thus, TREM-1 may be a potential diagnostic and therapeutic target in human kidney disease.