In view of the rapid preclinical development of cell-based therapies for neurodegenerative disorders, traumatic brain injury, and tumors, the safe and efficient delivery and targeting of therapeutic cells to the central nervous system is critical for maintaining therapeutic efficacy and safety in the respective disease models. Our previous data demonstrated therapeutically efficacious and targeted delivery of mesenchymal stem cells (MSCs) to the brain in the rat 6-hydroxydopamine model of Parkinson's disease (PD). The present study examined delivery of bone marrow-derived MSCs, macrophages, and microglia to the brain in a transgenic model of PD [(Thy1)-h[A30P] aS] and an APP/PS1 model of Alzheimer's disease (AD) via intranasal application (INA). INA of microglia in naive BL/6 mice led to targeted and effective delivery of cells to the brain. Quantitative PCR analysis of eGFP DNA showed that the brain contained the highest amount of eGFP-microglia (up to 2.1 × 104) after INA of 1 × 106 cells, while the total amount of cells detected in peripheral organs did not exceed 3.4 × 103. Seven days after INA, MSCs expressing eGFP were detected in the olfactory bulb (OB), cortex, amygdala, striatum, hippocampus, cerebellum, and brainstem of (Thy1)-h[A30P] aS transgenic mice, showing predominant distribution within the OB and brainstem. INA of eGFP-expressing macrophages in 13-month-old APP/PS1 mice led to delivery of cells to the OB, hippocampus, cortex, and cerebellum. Both MSCs and macrophages contained Iba-1-positive population of small microglia-like cells and Iba-1-negative large rounded cells showing either intracellular amyloid β (macrophages in APP/PS1 model) or α-synuclein [MSCs in (Thy1)-h[A30P] aS model] immunoreactivity. Here, we show, for the first time, intranasal delivery of cells to the brain of transgenic PD and AD mouse models. Additional work is needed to determine the optimal dosage (single treatment regimen or repeated administrations) to achieve functional improvement in these mouse models with intranasal microglia/macrophages and MSCs. This manuscript is published as part of the International Association of Neurorestoratology (IANR) special issue of Cell Transplantation.

Heterotrimeric G proteins of the Gαi family have been implicated in signaling pathways regulating cell migration in immune diseases. The Gαi-protein–coupled C5a receptor is a critical regulator of IgG FcR function in experimental models of immune complex (IC)–induced inflammation. By using mice deficient for Gαi2 or Gαi3, we show that Gαi2 is necessary for neutrophil influx in skin and lung Arthus reactions and agonist-induced neutrophilia in the peritoneum, whereas Gαi3 plays a less critical but variable role. Detailed analyses of the pulmonary IC-induced inflammatory response revealed several shared functions of Gαi2 and Gαi3, including mediating C5a anaphylatoxin receptor–induced activation of macrophages, involvement in alveolar production of chemokines, transition of neutrophils from bone marrow into blood, and modulation of CD11b and CD62L expression that account for neutrophil adhesion to endothelial cells. Interestingly, C5a-stimulated endothelial polymorphonuclear neutrophil transmigration, but not chemotaxis, is enhanced versus reduced in the absence of neutrophil Gαi3 or Gαi2, respectively, and knockdown of endothelial Gαi2 caused decreased transmigration of wild-type neutrophils. These data demonstrate that Gαi2 and Gαi3 contribute to inflammation by redundant, overlapping, and Gαi-isoform–specific mechanisms, with Gαi2 exhibiting unique functions in both neutrophils and endothelial cells that appear essential for polymorphonuclear neutrophil recruitment in IC disease.

Various heterotrimeric Gi proteins are considered to be involved in cell migration and effector function of immune cells. The underlying mechanisms, how they control the activation of myeloid effector cells, are not well understood. To elucidate isoform-redundant and -specific roles for Gαi proteins in these processes, we analyzed mice genetically deficient in Gαi2 or Gαi3. First, we show an altered distribution of tissue macrophages and blood monocytes in the absence of Gαi2 but not Gαi3. Gαi2-deficient but not wild-type or Gαi3-deficient mice exhibited reduced recruitment of macrophages in experimental models of thioglycollate-induced peritonitis and LPS-triggered lung injury. In contrast, genetic ablation of Gαi2 had no effect on Gαi-dependent peritoneal cytokine production in vitro and the phagocytosis-promoting function of the Gαi-coupled C5a anaphylatoxin receptor by liver macrophages in vivo. Interestingly, actin rearrangement and CCL2- and C5a anaphylatoxin receptor-induced chemotaxis but not macrophage CCR2 and C5a anaphylatoxin receptor expression were reduced in the specific absence of Gαi2. Furthermore, knockdown of Gαi2 caused decreased cell migration and motility of RAW 264.7 cells, which was rescued by transfection of Gαi2 but not Gαi3. These results indicate that Gαi2, albeit redundant to Gαi3 in some macrophage activation processes, clearly exhibits a Gαi isoform-specific role in the regulation of macrophage migration.

Dendritic cells (DCs) as professional antigen-presenting cells play an important role in the initiation and modulation of the adaptive immune response. However, their role in the innate immune response against bacterial infections is not completely defined. Here we have analyzed the role of DCs and their impact on the innate anti-bacterial host defense in an experimental infection model of Yersinia enterocolitica (Ye). We used CD11c-diphtheria toxin (DT) mice to deplete DCs prior to severe infection with Ye. DC depletion significantly increased animal survival after Ye infection. The bacterial load in the spleen of DC-depleted mice was significantly lower than that of control mice throughout the infection. DC depletion was accompanied by an increase in the serum levels of CXCL1, G-CSF, IL-1α, and CCL2 and an increase in the numbers of splenic phagocytes. Functionally, splenocytes from DC-depleted mice exhibited an increased bacterial killing capacity compared to splenocytes from control mice. Cellular studies further showed that this was due to an increased production of reactive oxygen species (ROS) by neutrophils. Adoptive transfer of neutrophils from DC-depleted mice into control mice prior to Ye infection reduced the bacterial load to the level of Ye-infected DC-depleted mice, suggesting that the increased number of phagocytes with additional ROS production account for the decreased bacterial load. Furthermore, after incubation with serum from DC-depleted mice splenocytes from control mice increased their bacterial killing capacity, most likely due to enhanced ROS production by neutrophils, indicating that serum factors from DC-depleted mice account for this effect. In summary, we could show that DC depletion triggers phagocyte accumulation in the spleen and enhances their anti-bacterial killing capacity upon bacterial infection.