Sophorolipid Butyl Ester Diacetate Does Not Affect Macrophage Polarization but Enhances Astrocytic Glial Fibrillary Acidic Protein Expression at Micromolar Concentrations in Vitro

Abstract

Peritoneal macrophages (PMACs) and spinal cord astrocytes were exposed to varying concentrations of soluble sophorolipid butyl ester diacetate (SLBEDA) in vitro. Macrophages and astrocytes demonstrated no decrease in viability in response to SLBEDA. Studying pro-and antiinflammatory genes, PMACs did not show a shift toward a pro-inflammatory phenotype. However, at higher concentrations (3 and 30 μM), astrocytes showed an increase in their expression of glial acidic fibrillary protein. This novel category of compounds poses low risk to PMAC and astrocyte viability; however, the effect on PMAC polarization and astrocyte reactivity requires more elucidation. Graphical abstract Supporting InformationThe Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acschemneuro.6b00451. Additional methods and results for SLBEDA synthesis and characterization and plasticity of PMAC polarization test (PDF) Notes Any opinions, findings, and conclusions or recommendations expressed in this material are those of the author(s) and do not necessarily reflect the views of the National Science Foundation. The authors declare no competing financial interest. (SCI). 1 The mechanical insult accompanying injury produces hemorrhaging, ischemia, and edema that result in an expanding lesion. 2 After approximately 2 h, the injury progresses to a secondary injury phase characterized by inflammation. Neutrophils first infiltrate the injury site followed by the activation of microglia and the recruitment of blood monocytes that differentiate and respond to the environment by polarizing to a pro-inflammatory or an antiinflammatory phenotype. M1, pro-inflammatory, macrophages are prevalent during the first several days postinjury 3 but persist weeks postinjury. 2,4 These macrophages produce high levels of reactive oxygen species (ROS) and pro-inflammatory cytokines. 3 Ultimately, this M1 macrophage pro-inflammation is neurotoxic and results in axonal dieback 3,4 and causes neuronal and glial necrosis. 5 In addition to macrophages, astrocytes also play an important role post-SCI. Astrocytes are a heterogeneous population of glial cells present in the central nervous system (CNS) and are primarily responsible for the overall maintenance of homeostasis in the CNS. 6 Following SCI, astrocytes at and around the site of the injury are activated, leading to the formation of a glial scar which acts as a chemical and physical barrier to regenerating axons. 7 The activation/reactivity of astrocytes is characterized by an increase in migration/proliferation 8,9 along with an increase in the production of the intermediate filament glial fibrillary acidic protein (GFAP) 10 and extracellular chondroitin sulfate proteoglycans (CSPGs). 11 GFAP synthesis is a meaningful aspect of the developmental plan of astrocyte differentiation, and it is a part of the reactive response to several CNS injuries. 12 After the injury, astrocytes show a decrease in GFAP until 1 day postinjury (dpi) followed by a steady increas...

Bone Marrow-Derived and Elicited Peritoneal Macrophages Are Not Created Equal: The Questions Asked Dictate the Cell Type Used

“…Quantitative polymerase chain reaction was conducted per previous lab protocols (39). The primers used are listed in Table 1.…”

Impact of Select Sophorolipid Derivatives on Macrophage Polarization and Viability

“…For example, one study recently concluded that 30 μM B did not affect LPSinduced mouse peritoneal macrophage viability, consistent with our results for this SL-ester. 45 Other studies have found significant cytotoxic effects of L on a variety of cell types, 61 including normal colonic epithelial cells and various colorectal cancer cell lines at 0.03 or 0.1 μM, respectively. Finally, Fu and colleagues found that L had greater toxicity than M when tested on human pancreatic carcinoma cells, 26 Beyond confirming some of these general viability trends, to our knowledge this study is the first to examine the impact of a range of different SL formulations, concentrations, and time points on macrophage viability.…”

“…To our knowledge, the immunomodulatory properties of L and B on the macrophage phenotype have been documented only in the contexts of sepsis 29,47,67 and spinal cord injury. 45 Bluth and colleagues showed that treatment with natural SLs (L makes up the highest proportion 23 ) significantly reduced nitric oxide production from LPS-activated RAW 264.7 cells, consistent with the reduction of pro-inflammatory markers IL-6 and TNF-α noted with L herein. 29 In contrast, the same authors observed an increase in IL-1β and TNF-α but no change in IL-6 gene expression in an LPS-activated rat alveolar macrophage cell line exposed to SLs for 36 h. Moreover, Ziemba and colleagues found that the treatment of peritoneal macrophages with B (3 nM to 30 μM for 24 h) did not affect the gene expression of either M1 markers (nitric oxide synthase and IL-12p40) or M2 markers (arginase-1 and IL-10).…”

“…29 In contrast, the same authors observed an increase in IL-1β and TNF-α but no change in IL-6 gene expression in an LPS-activated rat alveolar macrophage cell line exposed to SLs for 36 h. Moreover, Ziemba and colleagues found that the treatment of peritoneal macrophages with B (3 nM to 30 μM for 24 h) did not affect the gene expression of either M1 markers (nitric oxide synthase and IL-12p40) or M2 markers (arginase-1 and IL-10). 45 These results are different from the general reduction in the M1 profile and enhancement of the M2 profile noted here with B treatment. The above discrepancies for L and SLester B may potentially be explained by a variety of factors, including differences in (1) end-point assessments (i.e., gene vs protein expression and marker type) and (2) culture parameters (i.e., time and dosage).…”

“…26,45,46 The upper limits of concentrations employed herein for the synthetic SL-esters are generally higher than in any other study investigating these polymers, whereas the lower limits are comparable to those in similar studies performed with SL-esters. 45 The concentrations tested herein for L are relatively lower but consistent in magnitude compared to in vitro studies performed to support in vivo findings. 29,30,47 The use of the diacetylated lactonic form of L, rather than the ring-opened deacetylated form (i.e., acidic SL), was chosen for two reasons.…”