Increased Complement Component Research Articles

Comparison of the effects of hesperidin over 30 and 60-day intervals on rainbow trout: a potential biostimulant to promote growth, immunological and antioxidant responses, and disease resistance

Abstract The current research evaluated the effects of dietary hesperidin (HSP) on growth parameters, digestive enzyme activities, innate immune markers, and antioxidant responses in rainbow trout, Oncorhynchus mykiss after the 30- and 60-day feeding trial. Then, specimens were subjected to Yersinia ruckeri infection for 14 days. For this, six hundred rainbow trout juveniles (initial weight; 25.49±0.40 g) were fed with different levels of HSP including 0 (HSP0; control), 50 (HSP50), 100 (HSP100), 150 (HSP150), and 200 (HSP200) mg/kg feed. After 60 days, dietary HSP100 supplementation significantly improved growth and feed efficiency indices. The optimal dose of HSP based on the regression test for WG and FCR detected 122 and 131mg/kg, respectively. At the end of the 30th day, dietary HSP150 supplementation markedly boosted serum lysozyme (LYZ), myeloperoxidase (MPO), complement component C3, and immunoglobulin (Ig) levels, as decreased malondialdehyde (MDA) content. In the same period, dietary administration of HSP at the different concentrations markedly increased complement component C4 (HSP150 and HSP200), superoxide dismutase (SOD), skin mucus alkaline phosphatase (ALP) (HSP100- HSP200), skin mucus LYZ and catalase (CAT) (HSP100 and HSP 150), and skin mucus Ig level (HSP50- HSP200). After 60 days, all dietary HSP supplementation significantly improved lipase, serum C3, glutathione peroxidase (GPX), and skin mucus ALP, LYZ, and Ig levels, but decreased MDA, aspartate aminotransferase (AST), and alanine aminotransferase (ALT) values. After 60 days, dietary administration of HSP150 induced a significant discrepancy in the activity of serum LYZ, MPO, respiratory burst (RB), and lactate dehydrogenase (LDH) compared to the control group. In the same period, dietary HSP supplementation at different levels induced a remarkable increase in digestive protease activity (HSP100), serum ACH50 and CAT activities (HSP100 and HSP150), skin mucus peroxidase and serum SOD levels (HSP100-HSP200), and serum C4 value (HSP50, HSP150, and HSP200), but markedly decreased ALP activity (HSP150 and HSP200). In addition, dietary HSP supplementation, especially HSP150, markedly boosted disease resistance against Y. ruckeri infection. The current data demonstrated that the oral administration of 100 - 150 mg/kg HSP has a high potential to promote growth performance, disease resistance, and faster induction of immune and antioxidant responses.

Mitochondrial damage and clearance in retinal pigment epithelial cells.

Age-related macular degeneration (AMD) is a devastating eye disease that causes permanent vision loss in the central part of the retina, known as the macula. Patients with such severe visual loss face a reduced quality of life and are at a 1.5 times greater risk of death compared to the general population. Currently, there is no cure for or effective treatment for dry AMD. There are several mechanisms thought to underlie the disease, for example, ageing-associated chronic oxidative stress, mitochondrial damage, harmful protein aggregation and inflammation. As a way of gaining a better understanding of the molecular mechanisms behind AMD and thus developing new therapies, we have created a peroxisome proliferator-activated receptor gamma coactivator 1-alpha and nuclear factor erythroid 2-related factor 2 (PGC1α/NFE2L2) double-knockout (dKO) mouse model that mimics many of the clinical features of dry AMD, including elevated levels of oxidative stress markers, damaged mitochondria, accumulating lysosomal lipofuscin and extracellular drusen-like structures in retinal pigment epithelial cells (RPE). In addition, a human RPE cell-based model was established to examine the impact of non-functional intracellular clearance systems on inflammasome activation. In this study, we found that there was a disturbance in the autolysosomal machinery responsible for clearing mitochondria in the RPE cells of one-year-old PGC1α/NFE2L2-deficient mice. The confocal immunohistochemical analysis revealed an increase in autophagosome marker microtubule-associated proteins 1A/1B light chain 3B (LC3B) as well as multiple mitophagy markers such as PTE-induced putative kinase 1 (PINK1) and E3 ubiquitin ligase (PARKIN), along with signs of damaged mitochondria. However, no increase in autolysosome formation was detected, nor was there a colocalization of the lysosomal marker LAMP2 or the mitochondrial marker, ATP synthase β. There was an upregulation of late autolysosomal fusion Ras-related protein (Rab7) in the perinuclear space of RPE cells, together with autofluorescent aggregates. Additionally, we observed an increase in the numbers of Toll-like receptors 3 and 9, while those of NOD-like receptor 3 were decreased in PGC1α/NFE2L2 dKO retinal specimens compared to wild-type animals. There was a trend towards increased complement component C5a and increased involvement of the serine protease enzyme, thrombin, in enhancing the terminal pathway producing C5a, independent of C3. The levels of primary acute phase C-reactive protein and receptor for advanced glycation end products were also increased in the PGC1α/NFE2L2 dKO retina. Furthermore, selective proteasome inhibition with epoxomicin promoted both nicotinamide adenine dinucleotide phosphate (NADPH) oxidase and mitochondrial-mediated oxidative stress, leading to the release of mitochondrial DNA to the cytosol, resulting in potassium efflux-dependent activation of the absent in melanoma 2 (AIM2) inflammasome and the subsequent secretion of interleukin-1β in ARPE-19 cells. In conclusion, the data suggest that there is at least a relative decrease in mitophagy, increases in the amounts of C5 and thrombin and decreased C3 levels in this dry AMD-like model. Moreover, selective proteasome inhibition evoked mitochondrial damage and AIM2 inflammasome activation in ARPE-19 cells.

Yeast polysaccharide supplementation: impact on lactation, growth, immunity, and gut microbiota in Dezhou donkeys.

The Dezhou donkey, a prominent Chinese breed, is known for its remarkable size, rapid growth, and resilience to tough feeding conditions, and disease resistance. These traits are crucial in meeting the growing demand for Ejiao and donkey meat. Yeast polysaccharide (YPS), a functional polysaccharide complex known for its immune-enhancing and growth-promoting properties in livestock and poultry, remains relatively understudied in donkeys. This study aimed to investigate the impact of YPS supplementation on lactating and growing Dezhou donkey jennies and foals. Twelve 45-day-old Dezhou donkey foals and their jennies, matched for body weight and age, were randomly allocated to two dietary groups: a control group receiving a basal diet and an experimental group receiving the basal diet supplemented with 10 g/pen of YPS. The experiment was conducted over a 23-day period, during which donkey foals and lactating jennies were co-housed. The findings revealed that YPS supplementation had no adverse effects on milk production or composition in Dezhou donkey jennies but significantly increased feed intake. Additionally, YPS was associated with increased plasma glucose and creatinine concentrations in foals, while tending to decrease alkaline phosphatase, white blood cell count, red blood cell count, and hemoglobin levels (p < 0.10). Immune indices demonstrated that YPS supplementation elevated the levels of immunoglobulin A (IgA) and immunoglobulin G (IgG) in jennies (p < 0.05) and increased complement component C4 concentrations in foals (p < 0.05). Moreover, YPS positively influenced the fecal microbiome, promoting the abundance of beneficial microorganisms such as Lactobacillus and Prevotella in donkey foals and Terriporobacter and Cellulosilyticum in jennies, all of which contribute to enhanced feed digestion. Additionally, YPS induced alterations in the plasma metabolome for both jennies and foals, with a predominant presence of lipids and lipid-like molecules. Notably, YPS increased the concentrations of specific lipid metabolites, including 13,14-Dihydro PGF2a, 2-Isopropylmalic acid, 2,3-Dinor-TXB2, Triterpenoids, Taurocholic acid, and 3b-Allotetrahydrocortisol, all of which are associated with improved animal growth. In conclusion, this study suggests that dietary supplementation of YPS enhances feed intake, boosts immunity by increasing immunoglobulin levels, stimulates the growth-promoting gut microbiota (Lactobacillus and Prevotella), and exerts no adverse effects on the metabolism of both Dezhou donkey jennies and foals.

Activated endothelial cells induce a distinct type of astrocytic reactivity

Reactive astrogliosis is a universal response of astrocytes to abnormal events and injuries. Studies have shown that proinflammatory microglia can polarize astrocytes (designated A1 astrocytes) toward a neurotoxic phenotype characterized by increased Complement Component 3 (C3) expression. It is still unclear if inflammatory stimuli from other cell types may also be capable of inducing a subset of C3+ neurotoxic astrocytes. Here, we show that a subtype of C3+ neurotoxic astrocytes is induced by activated endothelial cells that is distinct from astrocytes activated by microglia. Furthermore, we show that endothelial-induced astrocytes have upregulated expression of A1 astrocytic genes and exhibit a distinctive extracellular matrix remodeling profile. Finally, we demonstrate that endothelial-induced astrocytes are Decorin-positive and are associated with vascular amyloid deposits but not parenchymal amyloid plaques in mouse models and AD/CAA patients. These findings demonstrate the existence of potentially extensive and subtle functional diversity of C3+-reactive astrocytes.

Type III TGF-β Receptor Down-Regulation Promoted Tumor Progression via Complement Component C5a Induction in Hepatocellular Carcinoma.

Simple SummaryThe clinical implications of TGFβR3 downregulation are currently unknown in hepatocellular carcinoma (HCC). Clinically, we identified that HCC patients with low expression levels of tumoral TGFβR3 exhibited significantly late tumor stages and shortened survival outcomes. Moreover, HCC patients developed lower plasma levels of TGFβR3 (sTGFβR3) (8.9 ng/mL) compared to healthy individuals (15.9 ng/mL), which represented a potential diagnostic marker. Similar to tumoral TGFβR3, low levels of plasma sTGFβR3 are also associated with poor clinical outcomes in HCC. To determine its tumor-suppressing capacities, continuous injection of sTGFβR3 in an orthotopic liver tumor model was performed, resulting in 2-fold tumor volume reduction compared to control. Decreased expression of TGFβR3 induced the upregulation of tumoral complement component C5a in HCC, which was found to contribute to poor clinical outcomes and promote tumor progression via a novel function in activating the tumor-promoting macrophages.Background and Aims—Transforming growth factor-beta (TGF-β) signaling orchestrates tumorigenesis and one of the family members, TGF-β receptor type III (TGFβR3), are distinctively under-expressed in numerous malignancies. Currently, the clinical impact of TGFβR3 down-regulation and the underlying mechanism remains unclear in hepatocellular carcinoma (HCC). Here, we aimed to identify the tumor-promoting roles of decreased TGFβR3 expression in HCC progression. Materials and Methods—For clinical analysis, plasma and liver specimens were collected from 100 HCC patients who underwent curative resection for the quantification of TGFβR3 by q-PCR and ELISA. To study the tumor-promoting mechanism of TGFβR3 downregulation, HCC mouse models and TGFβR3 knockout cell lines were applied. Results—Significant downregulation of TGFβR3 and its soluble form (sTGFβR3) were found in HCC tissues and plasma compared to healthy individuals (p < 0.01). Patients with <9.4 ng/mL sTGFβR3 exhibited advanced tumor stage, higher recurrence rate and shorter disease-free survival (p < 0.05). The tumor-suppressive function of sTGFβR3 was further revealed in an orthotopic mouse HCC model, resulting in 2-fold tumor volume reduction. In TGFβR3 knockout hepatocyte and HCC cells, increased complement component C5a was observed and strongly correlated with shorter survival and advanced tumor stage (p < 0.01). Interestingly, C5a activated the tumor-promoting Th-17 response in tumor associated macrophages. Conclusion—TGFβR3 suppressed tumor progression, and decreased expression resulted in poor prognosis in HCC patients through upregulation of tumor-promoting complement C5a.

C5a complement and cytokine signaling mediate the pronociceptive effects of complex regional pain syndrome patient IgM in fracture mice.

It has been proposed that complex regional pain syndrome (CRPS) is a posttraumatic autoimmune disease. Previously, we observed that B cells contribute to CRPS-like changes in a mouse tibia fracture model, and that early (<12 months duration) CRPS patient IgM antibodies have pronociceptive effects in the skin and spinal cord of muMT fracture mice lacking B cells. The current study evaluated the pronociceptive effects of intraplantar or intrathecal injections of early CRPS IgM (5 µg) in muMT fracture mice. Skin and lumbar spinal cord were collected for immunohistochemistry and polymerase chain reaction analyses. Wild-type mice exhibited postfracture increases in complement component C5a and its receptor expression in skin and spinal cord, predominantly on dermal macrophages and spinal microglia. Intraplantar IgM injection caused nociceptive sensitization in muMT fracture mice with increased complement component C1q and inflammatory cytokine expression, and these IgM effects were blocked by a C5a receptor antagonist (PMX53) or a global cytokine inhibitor (pentoxifylline). Intrathecal IgM injection also had pronociceptive effects with increased spinal cytokine expression, effects that were blocked by PMX53 or pentoxifylline treatment. Intrathecal injection of chronic (>12 months duration) CRPS patient IgM (but not IgG) caused nociceptive sensitization in muMT fracture mice, but intraplantar injection of chronic CRPS IgM or IgG had no effect. We postulate that CRPS IgM antibodies bind to neoantigens in the fracture limb skin and corresponding spinal cord to activate C5a complement signaling in macrophages and microglia, evoking proinflammatory cytokine expression contributing to nociceptive sensitization in the injured limb.

ANGPTL4 exacerbates pancreatitis by augmenting acinar cell injury through upregulation of C5a.

Pancreatitis is the inflammation of the pancreas. However, little is known about the genes associated with pancreatitis severity. Our microarray analysis of pancreatic tissues from mild and severe acute pancreatitis mice models identified angiopoietin‐like 4 (ANGPTL4) as one of the most significantly upregulated genes. Clinically, ANGPTL4 expression was also increased in the serum and pancreatic tissues of pancreatitis patients. The deficiency in ANGPTL4 in mice, either by gene deletion or neutralizing antibody, mitigated pancreatitis‐associated pathological outcomes. Conversely, exogenous ANGPTL4 exacerbated pancreatic injury with elevated cytokine levels and apoptotic cell death. High ANGPTL4 enhanced macrophage activation and infiltration into the pancreas, which increased complement component 5a (C5a) level through PI3K/AKT signaling. The activation of the C5a receptor led to hypercytokinemia that accelerated acinar cell damage and furthered pancreatitis. Indeed, C5a neutralizing antibody decreased inflammatory response in LPS‐activated macrophages and alleviated pancreatitis severity. In agreement, there was a significant positive correlation between C5a and ANGPTL4 levels in pancreatitis patients. Taken together, our study suggests that targeting ANGPTL4 is a potential strategy for the treatment of pancreatitis.

Quercetin improves immune function in Arbor Acre broilers through activation of NF-κB signaling pathway

Quercetin, the main component of flavonoids, has a wide range of biological actions. Quercetin can be made into a variety of additives for practice, because of the stable chemical structure and water-soluble derivatives. This study was intended to explore the effects of quercetin on immune function and its regulatory mechanism in Arbor Acre broiler to provide a practical basis for improving poultry immune function and figure out the optimum supplementation as functional feed additives. A total of 240 one-day-old healthy Arbor Acre broilers, similar in body weight, were randomly allotted to 4 treatments with 6 replicates, 10 broilers in each replicate and fed with diets containing quercetin at 0, 0.02, 0.04, and 0.06% for 6 wk. Blood and immune organs (spleen, thymus, and bursa) were collected from chickens at the end of the experiment. Growth performance, immune organs indexes, contents of serum immune molecules, splenic T lymphocyte proliferative responses, and expression of immune related genes were evaluated. The results showed that dietary quercetin had no significant effect (P > 0.05) on growth performance of broilers. Compared with control, 0.06% quercetin supplementation in diet significantly increased spleen index and thymus index (P < 0.05). It also increased the secretion of immune molecules including immunoglobulin A (IgA), interleukin-4 (IL-4) (P < 0.001), immunoglobulin M (IgM) (P = 0.007), complement component 4 (C4) (P = 0.001), and tumor necrosis factor-α (TNF-α) (P < 0.05). On the other hand, 0.02% quercetin supplementation significantly increased complement component 3 (C3) (P < 0.05). Additionally, both 0.04 and 0.06% quercetin supplementation significantly increased expression of TNF-α, TNF receptor associated factor-2 (TRAF-2), TNF receptor superfamily member 1B (TNFRSF1B), nuclear factor kappa-B p65 subunit (NF-κBp65), and interferon-γ (IFN-γ) mRNA (P < 0.05), and expression of NF-κB inhibitor-alpha (IκB-α) mRNA were significantly decreased (P < 0.05). Thus, quercetin improved immune function via NF-κB signaling pathway triggered by TNF-α.

Targeted Complement Inhibition at Synapses Prevents Microglial Synaptic Engulfment and Synapse Loss in Demyelinating Disease

Multiple sclerosis (MS) is a demyelinating, autoimmune disease of the central nervous system. While work has focused on myelin and axon loss in MS, less is known about mechanisms underlying synaptic changes. Using postmortem human MS tissue, a preclinical nonhuman primate model of MS, and two rodent models of demyelinating disease, we investigated synapse changes in the visual system. Similar to other neurodegenerative diseases, microglial synaptic engulfment and profound synapse loss were observed. In mice, synapse loss occurred independently of local demyelination and neuronal degeneration but coincided with gliosis and increased complement component C3, but not C1q, at synapses. Viral overexpression of the complement inhibitor Crry at C3-bound synapses decreased microglial engulfment of synapses and protected visual function. These results indicate that microglia eliminate synapses through the alternative complement cascade in demyelinating disease and identify a strategy to prevent synapse loss that may be broadly applicable to other neurodegenerative diseases. VIDEO ABSTRACT.

Decreased apolipoprotein A4 and increased complement component 3 as potential markers for papillary thyroid carcinoma: A proteomic study.

Thyroid carcinomas have comprised the fastest rising incidence of cancer in the past decade. Currently, the diagnosis of thyroid tumors is performed by the fine-needle aspiration biopsy (FNAB) method, which still holds some challenges and limitations, mostly in discriminating malignant and benign lesions. Therefore, the development of molecular markers to distinguish between these lesion types are in progress. A 2D-PAGE separation of proteins was performed followed by tandem mass spectrometry with the aim of discovering potential serum protein markers for papillary thyroid carcinoma and multinodular goiter. Protein-protein interaction network analysis revealed the most important pathways involved in the progression of papillary thyroid cancer. The enzyme-linked immunosorbent assay method was used to confirm a part of the results. The significantly altered proteins included C3, C4A, GC, HP, TTR, APOA4, APOH, ORM2, KRT10, AHSG, IGKV3-20, and IGKC. We also confirmed that increased complement component 3 and decreased apolipoprotein A4 occurred in papillary thyroid cancer. Network investigations demonstrated that complement activation cascades and PPAR signaling might play a role in the pathogenesis of thyroid cancer. The results demonstrated that serum proteomics could serve as a viable method for proposing novel potential markers for thyroid tumors. Surely, further research must be performed in larger cohorts to validate the results.

Resistance training prevents muscle fibrosis and atrophy via down-regulation of C1q-induced Wnt signaling in senescent mice.

Increased complement component 1q (C1q) secretion with aging leads to muscle fibrosis and atrophy whereas resistance training attenuates circulating C1q levels. This study aimed to clarify whether resistance exercise-induced reduction of C1q secretion contributes to the inhibition of fibrosis and atrophy in aged muscles. Young (13-wk-old) and aged (38-wk-old) senescence-accelerated mouse prone 1 mice were randomly assigned to one of 4 groups: a young or aged sedentary control group, or a young or aged resistance training (climbing a ladder 3 d/wk for 12 wk) group. We found that resistance training ameliorated muscle fibrosis and atrophy in aged mice, concomitant with decreased circulating and muscle C1q levels and attenuated activation of muscle Wnt signaling (glycogen synthase kinase β/β-catenin), including β-catenin in satellite (Pax7+/DAPI+) and fibroblast (vimentin+/DAPI+) cells. Furthermore, during muscle regeneration after mice were injured by cardiotoxin injection, we observed a reduction in circulating C1q levels, the inhibition of muscle fibrosis and repair, and decreased in the activation of muscle cytoplasmic and nuclear β-catenin in aged mice from the resistance training group, but these effects were cancelled by a single preadministration of exogenous recombinant C1q. In addition, resistance training attenuated aging-related muscle loss concomitant with decreased expression of both muscle ring-finger protein 1 and muscle atrophy F-box in the muscle. Thus, resistance training-induced changes in circulating C1q levels may contribute to the prevention of muscle fibrosis and atrophy via muscle Wnt signaling in senescent mice.-Horii, N., Uchida, M., Hasegawa, N., Fujie, S., Oyanagi, E., Yano, H., Hashimoto, T., Iemitsu, M. Resistance training prevents muscle fibrosis and atrophy via down-regulation of C1q-induced Wnt signaling in senescent mice.

Increase in complement iC3b is associated with anti-inflammatory cytokine expression during late pregnancy in mice.

Immunological tolerance between fetal allograft and mother is crucial for pregnancy establishment and maintenance; however, these mechanisms particularly those during the latter part of pregnancy have not been definitively elucidated. The aim of this study was to examine the presence and potential function of innate immunity characteristic to the middle to late pregnancy. We first characterized up-regulated proteins in decidua from day 11 pregnant (P11) mice using 2D-PAGE, followed by MALDI-TOF/MS analysis. These analyses identified increased complement component 3 (C3) and its derivatives in P11 decidua. We then found that in the decidual tissues, C3 mRNA increased on P15 and remained high on P19. C3 is converted to C3b and then iC3b by complement component factor I (Cfi) and complement receptor 1-like protein (Crry), both of which were present in P19 placentas. In addition, iC3b proteins and its receptor CR3 (Cd11b/Cd18) in decidual and placental tissues increased toward the latter phase of pregnancy. Moreover, CR3 subunit CD11b protein was predominantly localized to spongiotrophoblast layer in the P19 placenta. Because iC3b is known to induce anti-inflammatory cytokine production, the analysis was extended to examine changes in pro- and anti-inflammatory cytokines, Il12, Il10, and Tgfb1. Il12 expression decreased in P15 and P19 placenta, while high mRNA expression of Il10 and Tgfb1 was found in P19 placental tissues. Furthermore, placental Il10 and Tgfb1 mRNAs were down-regulated when pregnant mice were treated with an anti-C3 antibody, detecting C3, C3b and iC3b. These results indicated that C3 derivatives, in particular, iC3b and its receptor CR3 were up-regulated at the fetal-maternal interface, and suggest that iC3b may regulate the placental expression of anti-inflammatory cytokines, IL10 and TGFB1, during the latter phase of pregnancy.

Complement component 3 inhibition by an antioxidant is neuroprotective after cerebral ischemia and reperfusion in mice

Oxidative stress after stroke is associated with the inflammatory system activation in the brain. The complement cascade, especially the degradation products of complement component 3, is a key inflammatory mediator of cerebral ischemia. We have shown that pro-inflammatory complement component 3 is increased by oxidative stress after ischemic stroke in mice using DNA array. In this study, we investigated whether up-regulation of complement component 3 is directly related to oxidative stress after transient focal cerebral ischemia in mice and oxygen-glucose deprivation in brain cells. Persistent up-regulation of complement component 3 expression was reduced in copper/zinc-superoxide dismutase transgenic mice, and manganese-superoxide dismutase knock-out mice showed highly increased complement component 3 levels after transient focal cerebral ischemia. Antioxidant N-tert-butyl-α-phenylnitrone treatment suppressed complement component 3 expression after transient focal cerebral ischemia. Accumulation of complement component 3 in neurons and microglia was decreased by N-tert-butyl-α-phenylnitrone, which reduced infarct volume and impaired neurological deficiency after cerebral ischemia and reperfusion in mice. Small interfering RNA specific for complement component 3 transfection showed a significant increase in brain cells viability after oxygen-glucose deprivation. Our study suggests that the neuroprotective effect of antioxidants through complement component 3 suppression is a new strategy for potential therapeutic approaches in stroke.

Monitoring Molecular Changes Induced by Ischemia/Reperfusion in Human Free Muscle Flap Tissue Samples

Our current knowledge of the pathophysiological sequelae of ischemia or reperfusion (I/R) injury in free tissue transfer in reconstructive surgery is based on data obtained in animal experiments. In this study, we investigated the histologic and molecular changes after 11 free microsurgical muscle transfers in human muscle tissue. Biopsies of free muscle flap tissue were taken immediately before clipping of the pedicle and 5 days after ischemia and successful microanastomosis and restoration of the blood flow. Samples were analyzed histologically for edema formation and by immunohistochemistry for infiltration of inflammatory cells and angiogenesis. Expression levels of the inflammatory marker proteins interleukin-1β and tumor necrosis factor α and of complement component 3 as a major mediator of I/R injury were analyzed by real-time polymerase chain reaction. A TUNEL (terminal desoxynucleotidyl transferase-mediated-dUTP-nick-end-labeling) assay was used to assess apoptosis levels within the human muscle tissue. I/R injury leads to a significant up-regulation of inflammatory parameters, infiltration of inflammatory cells, and angiogenesis. Increased complement component 3 deposition and apoptosis of cells were accompanied by interstitial edema as indication for a pronounced postischemic inflammatory reaction within the muscle tissue after free tissue transfer. Our findings of molecular changes induced by I/R injury in human striated muscle tissue validate data obtained in animal models of I/R injury. The parameters and inflammatory patterns defined in this study will allow for the monitoring of the success of novel pharmaceutical strategies in the future and will help to transfer data obtained in animal work to the in vivo setting in human beings.

Overexpression of innate immune response genes in a model of recessive polycystic kidney disease

Defects in the primary cilium/basal body complex of renal tubular cells cause polycystic kidney disease (PKD). To uncover pathways associated with disease progression, we determined the kidney transcriptome of 10-day-old severely and mildly affected cpk mice, a model of recessive PKD. In the severe phenotype, the most highly expressed genes were those associated with the innate immune response including many macrophage markers, particularly those associated with a profibrotic alternative activation pathway. Additionally, gene expression of macrophage activators was dominated by the complement system factors including the central complement component 3. Additional studies confirmed increased complement component 3 protein levels in both cystic and non-cystic epithelia in the kidneys of cpk compared to wild-type mice. We also found elevated complement component 3 activation in two other mouse-recessive models and human-recessive PKD. Our results suggest that abnormal complement component 3 activation is a key element of progression in PKD.

Response-specific and ligand dose-dependent modulation of estrogen receptor (ER) alpha activity by ERbeta in the uterus.

Estrogen is of great importance in the regulation of uterine function. The aim of this study was to examine the individual physiological roles of each of the two receptors for estradiol, estrogen receptor (ER) alpha and ERbeta, and their potential comodulatory effects on gene expression and uterine growth using recently developed ER subtype-selective agonist ligands. The use of ER subtype-selective ligands provides an alternative, complementary approach to the use of receptor knockout mice. Administration of the ERalpha-selective ligand propyl pyrazole triol (PPT) to immature mice resulted in a significant increase in uterine weight, as well as bromodeoxyuridine incorporation and proliferating cell nuclear antigen expression in luminal epithelial cells. PPT also increased complement component 3, lactoferrin, and glucose-6-phosphate dehydrogenase (G6PDH), and decreased androgen receptor (AR) and progesterone receptor (PR) mRNA levels in uterine tissue, as did estradiol (E(2)). However, when compared with E(2), PPT was less effective in stimulating uterine weight, complement component 3, and G6PDH expression but was as effective as E(2) in regulating lactoferrin, AR, and PR expression. In contrast to the action of the ERalpha agonist PPT, the ERbeta agonist diarylpropionitrile (DPN) did not increase uterine weight or luminal epithelial cell proliferation at a dose that reduced G6PDH and elicited a decrease in PR and AR mRNA and protein expression. Interestingly, DPN reduced the uterine weight stimulation by PPT, and enhanced the effect of PPT in decreasing uterine PR and AR mRNA. These findings with ER subtype-selective ligands indicate that ERalpha is the major regulator of estrogen function in the uterus, but that ERbeta does exert effects on some uterine markers of estrogen action. In addition, ERbeta can modulate ERalpha activity in a response-specific and dose-dependent manner.