Engineered collagen-targeting therapeutics reverse lung and kidney fibrosis in mice

Fibrotic diseases are involved in 45% of deaths in the United States. In particular, fibrosis of the kidney and lung are major public health concerns due to their high prevalence and lack of existing treatment options. Here, we harness the pathophysiological features of fibrotic diseases, namely leaky vasculature and aberrant extracellular matrix (ECM) protein deposition (i.e. collagen), to target an anti-fibrotic biologic and a small molecule drug to disease sites of fibrosis, thus improving their therapeutic potential in mouse models of lung and kidney fibrosis. First, we identify and validate collagen-targeting drug delivery systems that preferentially accumulate in the diseased organs: von Willebrand Factor's A3 domain (VWF-A3) and decorin-derived collagen-binding peptide-conjugated micelles (CBP-micelles). We then engineer and recombinantly express novel candidate biologic therapies based on the anti-inflammatory cytokine IL-10: A3-IL-10 and A3-Serum Albumin-IL-10 (A3-SA-IL-10). Simultaneously, we stably encapsulate the potential anti-fibrotic water-insoluble drug, rapamycin, in CBP-micelles. We show that these novel formulations of therapeutics bind to collagen in vitro and that their efficacy in mouse models of lung and kidney fibrosis is improved, compared to free, untargeted drugs. Our results demonstrate that collagen-targeted anti-fibrotic drugs may be next generation therapies of high clinical potential.

Antioxidant and Inflammatory Effects of Nelumbo nucifera Gaertn. Leaves

This study is aimed at identifying the bioactive components in lotus leaf flavonoid extract (LLFE) and analyzing the antioxidant and anti-inflammatory activities of LLFE in vitro and in vivo. The flavonoids in LLFE were determined by UHPLC-MS/MS. The effect of LLFE on damaged 293T cells (H2O2, 0.3 mmol/L) was determined by MTT assay, and the activity of antioxidant enzymes was measured by kits. We studied the antioxidant and anti-inflammatory effects of LLFE on D-Gal/LPS (30 mg/kg·bw and 3 μg/kg·bw)-induced aging mice. We also evaluated the main organ index, pathological changes in the liver, lung, and kidney, liver function index, biochemical index, cytokine level, and mRNA expression level in serum and liver. The results showed that LLFE contains baicalein, kaempferol, kaempferid, quercetin, isorhamnetin, hyperoside, lespenephryl, and rutin. LLFE reduced the oxidative damage sustained by 293T cells, increased the levels of SOD, CAT, GSH, and GSH-Px, and decreased the level of MDA. The animal studies revealed that LLFE reduced oxidative damage and inflammation in injured mice, inhibited increases in AST, ALT, MDA, and NO, increased SOD, CAT, GSH, and GSH-Px levels, upregulated anti-inflammatory cytokines IL-10 and IL-12, and downregulated proinflammatory cytokines IL-6, IL-1β, TNF-α, and IFN-γ. Furthermore, the expression of antioxidant- and anti-inflammatory-related mRNA was consistent with the above results.

Manufacturing Multicolor LED-Based Phototherapy Device with a Novel 3D Design

Jaundice is a condition that results from an increase in bilirubin level in the blood. Its prevalence in newborns is around 60-70%. When this temporary jaundice becomes pathological and left untreated, significant damages may occur such as brain damage, vision loss, lung and kidney dysfunction, and even death. One of the methods used for the treatment of jaundice is phototherapy. In this study, a design has been made with 3 foldable LED panels to increase the target area. In addition, high-voltage LEDs with blue-green white wavelengths were used. Thus, it was aimed to minimize the risks of nausea and dizziness caused by intense blue light. An automatic system has been achieved by using temperature and light intensity sensors. The system will warn the user at temperatures and light intensity that are harmful to the baby.

A DNAmCULTURE Epigenetic Fingerprint Recapitulates Physiological Aging

Aging elicits dramatic changes to DNA methylation (DNAm), however the causes and consequences of such alterations to the epigenome remain unclear. The utility of biomarkers of aging based on DNAm patterns would be greatly enhanced if in vitro models existed that recapitulated physiological phenotypes such that modulation could garnish mechanistic insights. Using DNAm from serially passaged mouse embryonic fibroblasts, we developed a marker of culture aging and asked if culture phenotypes, like exhaustive replication, are epigenetically analogous to physiological aging. Our measure, termed DNAmCULTURE, accurately estimated passage number and was shown to strongly increase with age when examined in multiple tissues. Furthermore, we observed epigenetic alterations indicative of early cultured cells in animals undergoing caloric restriction and in lung and kidney fibroblasts re-programmed to iPSCs. This study identifies culture-derived alterations to the methylome as physiologically relevant and implicates culture aging as an important feature in known epigenetic aging phenomena.

HLA-G Is Widely Expressed by Mast Cells in Regions of Organ Fibrosis in the Liver, Lung and Kidney

We previously demonstrated that mast cells expressing HLA-G are associated with regions of hepatitis C virus-induced liver fibrosis. Here, we aimed to determine whether HLA-G expression in mast cells is specific to viral etiology, the liver, or to the general process of fibrosis. We enumerated HLA-G+ cells and mast cells by the immunohistochemistry of (i) liver blocks from 41 cases of alcoholic cirrhosis, (ii) 10 of idiopathic pulmonary fibrosis (IPF), and (iii) 10 of renal fibrosis. The nature of the HLA-G+ cells was specified by multiplex immunofluorescence using software. More than half of all HLA-G+ cells were mast cells in fibrotic areas of alcoholic cirrhosis and IPF. In the kidneys, subjected to fibrosis, the HLA-G+ cells were indeed mast cells but could not be counted. Moreover, in certain cases of the liver and lung, we observed a number of cellular nodes, which were secondary or tertiary follicles, in which HLA-G was highly expressed by B lymphocytes. In conclusion, HLA-G+ mast cells could be observed in the fibrotic regions of all organs studied. Previous studies suggest a protective role for HLA-G+ mast cells against inflammation and fibrosis. The observed follicles with B lymphocytes that express HLA-G may also reinforce their antifibrotic role.

Mds1 CreERT2-Based Lineage-Tracing Reveals Increasing Contributions of HSCs to Fetal Hematopoiesis and to Adult Tissue-Resident Macrophages in the Marrow

The ontogeny of the hematopoietic system consists of two broad programs. The first, an HSC-independent program, consists of overlapping waves of primitive, erythro-myeloid (EMP), and some lymphoid progenitors. HSC-independent hematopoiesis is required for normal fetal development, and provides self-renewing tissue-resident macrophage populations that persist in the adult. This is followed by the emergence of an HSC-dependent program that arises from arterial vessels within the body of the embryo. The overlapping emergence and lineage output of HSC-independent and HSC-derived hematopoiesis raises important questions regarding the identity and potential functional differences of their mature progeny. However, the transition from HSC-independent to HSC-derived hematopoiesis in the murine fetus remains incompletely characterized, particularly since the maturing erythroid, megakaryocytic and myeloid progeny of EMP and HSCs are currently not easily distinguishable. Additionally, lineage-tracing approaches have been challenging because they have relied largely on the temporal induction of promoters that are expressed both in HSC-independent progenitors and in HSCs, which have significant temporal overlap in their developmental emergence and result in incomplete or in mixed labeling. To help resolve this question, we have developed Mds1 CreERT2 mice, utilizing the first transcription start site of MECOM gene, which is expressed in HSC and emerging HSC (Yuasa et al., 2005 EMBO; Hou et al. 2020 Cell Research; Zhu et al. 2020, Blood). When mated with Rosa-YFP reporter mice and induced at E9.5 with tamoxifen, this construct lineage-traces pre-HSCs present in the E11.5 AGM region, as well as HSCs in the fetal liver and adult marrow. Importantly, no labeling of primitive erythroid cells, primitive macrophage-derived microglia, EMP, or EMP-derived cells in the E11.5 or E12.5 fetal liver was detected with tamoxifen induction at either E9.5 or E8.5. Analysis of E9.5 tamoxifen-treated Mds1 CreERT2Rosa26 LSL-YFP embryos indicates that HSCs have begun to generate small numbers of differentiating erythroid, myeloid and lymphoid progeny in the liver between E12.5 and E14.5. By E16.5, a significant proportion of differentiating erythroid, myeloid and B-cell lineage cells in the liver are HSC-derived, and HSCs have now begun to contribute erythroid and myeloid cells to the rapidly expanding pool of circulating blood cells. In the adult, we found increasing contributions of HSCs to macrophages in liver, lung and kidney. Interestingly, the majority of F4/80+ cells in the adult bone marrow and spleen were also lineage-traced in these mice. Thus, HSCs ultimately provide the majority of adult marrow macrophages that go on to self-maintain in the adult marrow (Hashimoto et al., 2013, Immunity). The Mds1 CreERT2 mouse model will serve as a useful to deconvolute the complexity of hematopoiesis as it unfolds in the embryo and functions postnatally. Disclosures Palis: Rubius Therapeutics: Consultancy.

First Report of Sarcocystis Masoni in a Captive Alpaca (Vicugna Pacos) From China

Background: Sarcocystosis is a parasitic disease caused by intracellular protozoan parasite of the genus Sarcocystis. Tissue samples of alpacas (n = 4) from Henan province (China) were screened for Sarcocystis spp. infection by histological examination, pepsin digestion, and molecular assays.Results:Sarcocystis spp. was detected in heart, liver, spleen, lung, and kidney of an alpaca by molecular assays. Many sarcocysts with inflammation responses were observed in this alpaca myocardium, and they showed a high similarity to Sarcocystis masoni by sequence analysis.Conclusion: This study is the first to demonstrate Sarcocystis spp. infection in alpaca from China. The higher parasite load in the alpaca myocardium indicated that it had contact with an environment contaminated with sporocysts, and that the alpaca was susceptible to Sarcocystis spp.