A Novel Approach for Meniscal Regeneration Using Kartogenin-Treated Autologous Tendon Graft

2017 ◽  
Vol 45 (14) ◽  
pp. 3289-3297 ◽  
Author(s):  
He Huang ◽  
Hongyao Xu ◽  
Jianning Zhao
Keyword(s):  
Chondrogenic Differentiation ◽  
Collagen Type ◽  
Tendon Graft ◽  
Histochemical Staining ◽  
Type I ◽  
Type Ii ◽  
Qrt Pcr ◽  
Autologous Tendon Graft

Background: The meniscus is one of the most commonly injured parts of the body, and meniscal healing is difficult. Hypothesis: Kartogenin (KGN) induces tendon stem cells (TSCs) to differentiate into cartilage cells in vitro and form meniscus-like tissue in vivo. A damaged meniscus can be replaced with a KGN-treated autologous tendon graft. Study Design: Controlled laboratory study. Methods: In the in vitro experiments, TSCs were isolated from rabbit patellar tendons and cultured with various concentrations of KGN, from 0 to 1000 µM. The effect of KGN on the chondrogenesis of TSCs in vitro was investigated by histochemical staining and quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR). The in vivo experiments were carried out on 6 New Zealand White rabbits by removing a meniscus from the rabbit knee and implanting an autologous tendon graft treated with KGN or saline. The meniscus formation in vivo was examined by histological analysis and immune staining. Results: The proliferation of TSCs was promoted by KGN in a concentration-dependent manner. Both histochemical staining and qRT-PCR showed that the chondrogenic differentiation of TSCs was increased with KGN concentration. After 3 months of implantation, the tendon graft treated with KGN formed a meniscus-like tissue with a white and glistening appearance, while the saline-treated tendon graft retained tendon-like tissue and appeared yellowish and unhealthy. Histochemical staining showed that after 3 months of implantation, the KGN-treated tendon graft had a structure similar to that of normal meniscus. Many cartilage-like cells and fibrocartilage-like tissues were found in the KGN-treated tendon graft. However, no cartilage-like cells were found in the saline-treated tendon graft after 3 months of implantation. Furthermore, the KGN-treated tendon graft was positively stained by both anti–collagen type I and type II antibodies, but the saline-treated tendon graft was not stained by collagen type II. Conclusion: The findings indicated that KGN can induce the differentiation of TSCs into cartilage-like cells in vitro and in vivo. The results suggest that KGN-treated tendon graft may be a good substitute for meniscal repair and regeneration. Clinical Relevance: This study revealed the direct effects of KGN on the chondrogenic differentiation of TSCs in vitro and in vivo. A KGN-treated autologous tendon graft induced formation of a meniscus-like tissue in vivo. This study provides a new cartilage regenerating technology for the treatment of damaged meniscus.

scholarly journals Different Phenotypes and Chondrogenic Responses of Human Menstrual Blood and Bone Marrow Mesenchymal Stem Cells to activin A and TGF-β3

2020 ◽  
Author(s):  
Ilona Uzieliene ◽  
Edvardas Bagdonas ◽  
Kazuto Hoshi ◽  
Tomoaki Sakamoto ◽  
Atsuhiko Hikita ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Chondrogenic Differentiation ◽  
Collagen Type ◽  
Activin A ◽  
Cell Surface Markers ◽  
Type Ii ◽  
Marrow Mesenchymal Stem Cells

Abstract Background: Due to its low capacity for self-repair, articular cartilage is highly susceptible to damage and deterioration, which leads to the development of degenerative joint diseases such as osteoarthritis. Menstrual blood-derived mesenchymal stem cells (MenSCs) are much less characterized compared to bone marrow mesenchymal stem cells (BMMSCs). However, MenSCs seem an attractive alternative to classical BMMSCs due to ease of access and broader differentiation capacity. The aim of this study was to evaluate chondrogenic differentiation potential of MenSCs and BMMSCs stimulated with transforming growth factor β (TGF-β3) and activin A, member of the TGF-β superfamily of proteins.Methods: MenSCs (n=6) and BMMSCs (n=5) were isolated from different healthy donors. Expression of cell surface markers CD90, CD73, CD105, CD44, CD45, CD14, CD36, CD55, CD54, CD63, CD106, CD34, CD10, Notch1 was analysed by flow cytometry. Cell proliferation capacity was determined using CCK-8 proliferation kit. Adipogenic differentiation capacity was evaluated according to Oil-Red staining, osteogenic differentiation - Alizarin Red staining. Chondrogenic differentiation (Activin A and TGF-β3 stimulation) was induced in vitro and in vivo (subcutaneous scaffolds in nude BALB/c mice) and investigated by histologically and by expression of chondrogenic genes (collagen type II, aggrecan). Activin A protein production was evaluated by ELISA.Results: MenSCs exhibited a higher proliferation rate, as compared to BMMSCs, and a different expression profile of several cell surface markers. Activin A stimulated collagen type II gene expression and glycosaminoglycan synthesis in TGF-β3 treated MenSCs but not in BMMSCs, both in vitro and in vivo, although the effects of TGF-β3 alone were more pronounced in BMMSCs in vitro. Conclusion: These data suggest that activin A exerts differential effects on the induction of chondrogenic differentiation in MenSCs vs. BMMSCs, which implies that different mechanisms of chondrogenic regulation are activated in these cells. Following further optimisation of differentiation protocols and the choice of growth factors, potentially including activin A, MenSCs may turn out to be a promising population of stem cells for the development of cell-based therapies with the capacity to stimulate cartilage repair and regeneration.Trial registration: Not applicable.

scholarly journals Hydrogel-hydroxyapatite-monomeric collagen type-I scaffold with low-frequency electromagnetic field treatment enhances osteochondral repair in rabbits

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Jiyuan Yan ◽  
Chaoxu Liu ◽  
Chang Tu ◽  
Ruizhuo Zhang ◽  
Xiangyu Tang ◽  
...  
Keyword(s):  
Chondrogenic Differentiation ◽  
Collagen Type ◽  
Composite Scaffold ◽  
Low Frequency ◽  
Collagen Type I ◽  
Type I ◽  
Osteochondral Defects ◽  
Osteochondral Repair

Abstract Background Cartilage damage is a common medical issue in clinical practice. Complete cartilage repair remains a significant challenge owing to the inferior quality of regenerative tissue. Safe and non-invasive magnetic therapy combined with tissue engineering to repair cartilage may be a promising breakthrough. Methods In this study, a composite scaffold made of Hydroxyapatite-Collagen type-I (HAC) and PLGA-PEG-PLGA thermogel was produced to match the cartilage and subchondral layers in osteochondral defects, respectively. Bone marrow mesenchymal stem cells (BMSC) encapsulated in the thermogel were stimulated by an electromagnetic field (EMF). Effect of EMF on the proliferation and chondrogenic differentiation potential was evaluated in vitro. 4 mm femoral condyle defect was constructed in rabbits. The scaffolds loaded with BMSCs were implanted into the defects with or without EMF treatment. Effects of the combination treatment of the EMF and composite scaffold on rabbit osteochondral defect was detected in vivo. Results In vitro experiments showed that EMF could promote proliferation and chondrogenic differentiation of BMSCs partly by activating the PI3K/AKT/mTOR and Wnt1/LRP6/β-catenin signaling pathway. In vivo results further confirmed that the scaffold with EMF enhances the repair of osteochondral defects in rabbits, and, in particular, cartilage repair. Conclusion Hydrogel-Hydroxyapatite-Monomeric Collagen type-I scaffold with low-frequency EMF treatment has the potential to enhance osteochondral repair.

scholarly journals Different phenotypes and chondrogenic responses of human menstrual blood and bone marrow mesenchymal stem cells to activin A and TGF-β3

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Ilona Uzieliene ◽  
Edvardas Bagdonas ◽  
Kazuto Hoshi ◽  
Tomoaki Sakamoto ◽  
Atsuhiko Hikita ◽  
...  
Keyword(s):  
Stromal Cells ◽  
Chondrogenic Differentiation ◽  
Collagen Type ◽  
Activin A ◽  
Surface Markers ◽  
Cell Surface Markers ◽  
Type Ii ◽  
Collagen Type Ii

Abstract Background Due to its low capacity for self-repair, articular cartilage is highly susceptible to damage and deterioration, which leads to the development of degenerative joint diseases such as osteoarthritis (OA). Menstrual blood-derived mesenchymal stem/stromal cells (MenSCs) are much less characterized, as compared to bone marrow mesenchymal stem/stromal cells (BMMSCs). However, MenSCs seem an attractive alternative to classical BMMSCs due to ease of access and broader differentiation capacity. The aim of this study was to evaluate chondrogenic differentiation potential of MenSCs and BMMSCs stimulated with transforming growth factor β (TGF-β3) and activin A. Methods MenSCs (n = 6) and BMMSCs (n = 5) were isolated from different healthy donors. Expression of cell surface markers CD90, CD73, CD105, CD44, CD45, CD14, CD36, CD55, CD54, CD63, CD106, CD34, CD10, and Notch1 was analyzed by flow cytometry. Cell proliferation capacity was determined using CCK-8 proliferation kit and cell migration ability was evaluated by scratch assay. Adipogenic differentiation capacity was evaluated according to Oil-Red staining and osteogenic differentiation according to Alizarin Red staining. Chondrogenic differentiation (activin A and TGF-β3 stimulation) was investigated in vitro and in vivo (subcutaneous scaffolds in nude BALB/c mice) by expression of chondrogenic genes (collagen type II, aggrecan), GAG assay and histologically. Activin A protein production was evaluated by ELISA during chondrogenic differentiation in monolayer culture. Results MenSCs exhibited a higher proliferation rate, as compared to BMMSCs, and a different expression profile of several cell surface markers. Activin A stimulated collagen type II gene expression and glycosaminoglycan synthesis in TGF-β3 treated MenSCs but not in BMMSCs, both in vitro and in vivo, although the effects of TGF-β3 alone were more pronounced in BMMSCs in vitro. Conclusion These data suggest that activin A exerts differential effects on the induction of chondrogenic differentiation in MenSCs vs. BMMSCs, which implies that different mechanisms of chondrogenic regulation are activated in these cells. Following further optimization of differentiation protocols and the choice of growth factors, potentially including activin A, MenSCs may turn out to be a promising population of stem cells for the development of cell-based therapies with the capacity to stimulate cartilage repair and regeneration in OA and related osteoarticular disorders.

scholarly journals Plant Recombinant Human Collagen Type I Hydrogels for Corneal Regeneration

2021 ◽  
Author(s):  
Michel Haagdorens ◽  
Elle Edin ◽  
Per Fagerholm ◽  
Marc Groleau ◽  
Zvi Shtein ◽  
...  
Keyword(s):  
Collagen Type ◽  
Collagen Type I ◽  
Collagen Fibrils ◽  
Type I ◽  
Safe Alternative ◽  
Human Donor ◽  
Human Collagen ◽  
Donor Corneas

Abstract Purpose To determine feasibility of plant-derived recombinant human collagen type I (RHCI) for use in corneal regenerative implants Methods RHCI was crosslinked with 1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide (EDC) and N-hydroxysuccinimide (NHS) to form hydrogels. Application of shear force to liquid crystalline RHCI aligned the collagen fibrils. Both aligned and random hydrogels were evaluated for mechanical and optical properties, as well as in vitro biocompatibility. Further evaluation was performed in vivo by subcutaneous implantation in rats and corneal implantation in Göttingen minipigs. Results Spontaneous crosslinking of randomly aligned RHCI (rRHCI) formed robust, transparent hydrogels that were sufficient for implantation. Aligning the RHCI (aRHCI) resulted in thicker collagen fibrils forming an opaque hydrogel with insufficient transverse mechanical strength for surgical manipulation. rRHCI showed minimal inflammation when implanted subcutaneously in rats. The corneal implants in minipigs showed that rRHCI hydrogels promoted regeneration of corneal epithelium, stroma, and nerves; some myofibroblasts were seen in the regenerated neo-corneas. Conclusion Plant-derived RHCI was used to fabricate a hydrogel that is transparent, mechanically stable, and biocompatible when grafted as corneal implants in minipigs. Plant-derived collagen is determined to be a safe alternative to allografts, animal collagens, or yeast-derived recombinant human collagen for tissue engineering applications. The main advantage is that unlike donor corneas or yeast-produced collagen, the RHCI supply is potentially unlimited due to the high yields of this production method. Lay Summary A severe shortage of human-donor corneas for transplantation has led scientists to develop synthetic alternatives. Here, recombinant human collagen type I made of tobacco plants through genetic engineering was tested for use in making corneal implants. We made strong, transparent hydrogels that were tested by implanting subcutaneously in rats and in the corneas of minipigs. We showed that the plant collagen was biocompatible and was able to stably regenerate the corneas of minipigs comparable to yeast-produced recombinant collagen that we previously tested in clinical trials. The advantage of the plant collagen is that the supply is potentially limitless.

scholarly journals WNT-5A regulates TGF-β-related activities in liver fibrosis

2017 ◽  
Vol 312 (3) ◽  
pp. G219-G227 ◽  
Author(s):  
Leonie Beljaars ◽  
Sara Daliri ◽  
Christa Dijkhuizen ◽  
Klaas Poelstra ◽  
Reinoud Gosens
Keyword(s):  
Liver Fibrosis ◽  
Functional Role ◽  
Collagen Type ◽  
Collagen Type I ◽  
Matrix Proteins ◽  
Type I ◽  
Human Livers

WNT-5A is a secreted growth factor that belongs to the noncanonical members of the Wingless-related MMTV-integration family. Previous studies pointed to a connection between WNT-5A and the fibrogenic factor TGF-β warranting further studies into the functional role of WNT-5A in liver fibrosis. Therefore, we studied WNT-5A expressions in mouse and human fibrotic livers and examined the relation between WNT-5A and various fibrosis-associated growth factors, cytokines, and extracellular matrix proteins. WNT-5A gene and protein expressions were significantly increased in fibrotic mouse and human livers compared with healthy livers. Regression or therapeutic intervention in mice resulted in decreased hepatic WNT-5A levels paralleled by lower collagen levels. Immunohistochemical analysis showed WNT-5A staining in fibrotic septa colocalizing with desmin staining indicating WNT-5A expression in myofibroblasts. In vitro studies confirmed WNT-5A expression in this cell type and showed that TGF-β significantly enhanced WNT-5A expression in contrast to PDGF-BB and proinflammatory cytokines IL-1β and TNF-α. Additionally, TGF-β induces the expression of the WNT receptors FZD2 and FZD8. After silencing of WNT-5A, reduced levels of collagen type I, vimentin, and fibronectin in TGF-β-stimulated myofibroblasts were measured compared with nonsilencing siRNA-treated controls. Interestingly, the antifibrotic cytokine IFNγ suppressed WNT-5A in vitro and in vivo. IFNγ-treated fibrotic mice showed significantly less WNT-5A expression compared with untreated fibrotic mice. In conclusion, WNT-5A paralleled collagen I levels in fibrotic mouse and human livers. WNT-5A expression in myofibroblasts is induced by the profibrotic factor TGF-β and plays an important role in TGF-β-induced regulation of fibrotic matrix proteins, whereas its expression can be reversed upon treatment, both in vitro and in vivo. NEW & NOTEWORTHY This study describes the localization and functional role of WNT-5A in human and mouse fibrotic livers. Hepatic WNT-5A expression parallels collagen type I expression. In vivo and in vitro, the myofibroblasts were identified as the key hepatic cells producing WNT-5A. WNT-5A is under control of TGF-β and its activities are primarily profibrotic.

scholarly journals Biological Relevance of Colony Morphology and Phenotypic Switching by Burkholderia pseudomallei

2006 ◽  
Vol 189 (3) ◽  
pp. 807-817 ◽  
Author(s):  
Narisara Chantratita ◽  
Vanaporn Wuthiekanun ◽  
Khaemaporn Boonbumrung ◽  
Rachaneeporn Tiyawisutsri ◽  
Mongkol Vesaratchavest ◽  
...  
Keyword(s):  
Burkholderia Pseudomallei ◽  
Vaccine Development ◽  
Colony Morphology ◽  
Phenotypic Switching ◽  
Type I ◽  
Type Ii ◽  
Altered Expression ◽  
Replication Fitness

ABSTRACT Melioidosis is a notoriously protracted illness and is difficult to cure. We hypothesize that the causative organism, Burkholderia pseudomallei, undergoes a process of adaptation involving altered expression of surface determinants which facilitates persistence in vivo and that this is reflected by changes in colony morphology. A colony morphotyping scheme and typing algorithm were developed using clinical B. pseudomallei isolates. Morphotypes were divided into seven types (denoted I to VII). Type I gave rise to other morphotypes (most commonly type II or III) by a process of switching in response to environmental stress, including starvation, iron limitation, and growth at 42°C. Switching was associated with complex shifts in phenotype, one of which (type I to type II) was associated with a marked increase in production of factors putatively associated with in vivo concealment. Isogenic types II and III, derived from type I, were examined using several experimental models. Switching between isogenic morphotypes occurred in a mouse model, where type II appeared to become adapted for persistence in a low-virulence state. Isogenic type II demonstrated a significant increase in intracellular replication fitness compared with parental type I after uptake by epithelial cells in vitro. Isogenic type III demonstrated a higher replication fitness following uptake by macrophages in vitro, which was associated with a switch to type II. Mixed B. pseudomallei morphologies were common in individual clinical specimens and were significantly more frequent in samples of blood, pus, and respiratory secretions than in urine and surface swabs. These findings have major implications for therapeutics and vaccine development.

scholarly journals LeuRS can leucylate type I and type II tRNALeus in Streptomyces coelicolor

2019 ◽  
Vol 47 (12) ◽  
pp. 6369-6385
Author(s):  
Jia-Yi Fan ◽  
Qian Huang ◽  
Quan-Quan Ji ◽  
En-Duo Wang
Keyword(s):  
Tertiary Structure ◽  
Streptomyces Coelicolor ◽  
Catalytic Efficiency ◽  
Type I ◽  
Type Ii ◽  
Specific Domain ◽  
Recognition Mechanism ◽  
Variable Loop

Abstract Transfer RNAs (tRNAs) are divided into two types, type I with a short variable loop and type II with a long variable loop. Aminoacylation of type I or type II tRNALeu is catalyzed by their cognate leucyl-tRNA synthetases (LeuRSs). However, in Streptomyces coelicolor, there are two types of tRNALeu and only one LeuRS (ScoLeuRS). We found that the enzyme could leucylate both types of ScotRNALeu, and had a higher catalytic efficiency for type II ScotRNALeu(UAA) than for type I ScotRNALeu(CAA). The results from tRNA and enzyme mutagenesis showed that ScoLeuRS did not interact with the canonical discriminator A73. The number of nucleotides, rather than the type of base of the variable loop in the two types of ScotRNALeus, was determined as important for aminoacylation. In vitro and in vivo assays showed that the tertiary structure formed by the D-loop and TψC-loop is more important for ScotRNALeu(UAA). We showed that the leucine-specific domain (LSD) of ScoLeuRS could help LeuRS, which originally only leucylates type II tRNALeu, to aminoacylate type I ScotRNALeu(CAA) and identified the crucial amino acid residues at the C-terminus of the LSD to recognize type I ScotRNALeu(CAA). Overall, our findings identified a rare recognition mechanism of LeuRS to tRNALeu.

External PS in Sickle RBC: Relationships among Aminophospholipid Translocase (APLT), Phospholipid Scramblase (PLSCR), Cell Maturity, and Cell Density.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 148-148
Author(s):  
Latorya E. Arnold ◽  
Mary B. Palascak ◽  
Clinton H. Joiner ◽  
Robert S. Franco
Keyword(s):  
Annexin V ◽  
Type I ◽  
Type Ii ◽  
Color Flow ◽  
Low Level ◽  
Aminophospholipid Translocase ◽  
Phospholipid Scramblase ◽  
High Level

Abstract External phosphatidylserine (PS) is present on some sickle RBC and may contribute to thrombogenesis, endothelial adhesion, and shortened RBC lifespan. Phospholipid scramblase (PLSCR) disrupts phospholipid (PL) asymmetry by causing nonspecific PL equilibration across the membrane. Aminophospholipid translocase (APLT) maintains PL asymmetry by returning externalized PS to the inner membrane leaflet. It has been proposed that both APLT inhibition and PLSCR activation are required for PS externalization. Sickle RBC with low level external PS (Type I PS+) are present in cells of all densities and include some reticulocytes. Sickle RBC with high external PS (Type II PS+) are primarily found in the dense fraction. Type II cells are thought to be more important because: the high level of external PS should have greater consequence; high level external PS occurs primarily in pathologically dehydrated sickle RBC; and low level external PS appears to be physiological in immature RBC. We have previously shown that dense, dehydrated sickle RBC, including the small number of dense transferrin receptor positive (TfR+) reticulocytes, have markedly inhibited APLT. In the current studies, we examined the relationships among external PS, APLT, PLSCR, and density in mature RBC and TfR+ reticulocytes using 3-color flow cytometry. APLT and PLSCR activities were assayed using fluorescent PL analogues (NBD-PS and NBD-PC, respectively), and expressed as the fraction of probe internalized. External PS was measured with Annexin V-PE and TfR+ reticulocytes were identified with anti-TfR-PE/Cy5. PS+ cells had lower APLT activity compared to PS- cells that did not reach significance for n=3 (NBD-PS internalization fraction for PS-: 0.586±0.053; Type I PS+: 0.517±0.158, Type II PS+: 0.523±0.033). PS- sickle RBC had a uniformly low PLSCR activity similar to normal RBC (NBD-PC internalization fractions ∼ 0.1). In mature sickle RBC, PLSCR was more active in PS+ cells (PS-: 0.097±0.096; Type I PS+: 0.163±0.070, Type II PS+: 0.248±0.043; n=3; PS- vs Type I PS+: p=0.06; PS- vs Type II PS+: p=0.04; Type I versus Type II: p=0.03). TfR+ reticulocytes had increased APLT and PLSCR activity compared to mature sickle RBC, but there was no apparent relationship between PLSCR and external PS. Since dense sickle RBC had markedly inhibited APLT, we evaluated the relationship between dehydration and APLT activity. Dehydration of AA RBC from an MCHC of 35.6±2.2 to 49.2±2.0 g/dL inhibited APLT (from 0.484±0.068 to 0.301±0.076; n=7, p= 0.01). Dehydration of SS RBC from an MCHC of 34.8±3.5 to 50.1±3.9 g/dL also inhibited APLT (from 0.460±0.060 to 0.361±0.047; n=3, p=0.006), but not as low as in SS RBC dehydrated in vivo (0.222±0.036 at 44.7±5.6 g/dL; n=4, p=0.007 vs. SS RBC dehydrated in vitro). Rehydration of AA and SS RBC that had been dehydrated in vitro reversed APLT inhibition. However, APLT activity was not reversed upon rehydration of sickle RBC dehydrated in vivo. In summary, our data show that: many dense sickle RBC with significantly inhibited APLT are PS-, indicating that APLT inhibition alone does not result in PS externalization; dehydration contributes to, but is not entirely responsible for, the APLT inhibition seen in dense sickle RBC; and PS+ sickle RBC have increased PLSCR activity.

scholarly journals Adjuvant properties of a biocompatible thermo-responsive polymer of N -isopropylacrylamide in autoimmunity and arthritis

2011 ◽  
Vol 8 (65) ◽  
pp. 1748-1759 ◽  
Author(s):  
Akhilesh Kumar Shakya ◽  
Ashok Kumar ◽  
Kutty Selva Nandakumar
Keyword(s):  
Antibody Response ◽  
Collagen Type ◽  
Physical Adsorption ◽  
Type Ii ◽  
Specific Igg ◽  
Ifn Γ ◽  
First Time ◽  
Molecular Weight Form

To evaluate the thermo-responsive poly( N -isopropylacrylamide) (PNiPAAm) polymer as an adjuvant, we synthesized PNiPAAm through free radical polymerization and characterized it both in vitro and in vivo . The polymer when mixed with collagen type II (CII) induced antigen-specific autoimmunity and arthritis. Mice immunized with PNiPAAm–CII developed significant levels of CII-specific IgG response comprising major IgG subclasses. Antigen-specific cellular recall response was also enhanced in these mice, while negligible level of IFN-γ was detected in splenocyte cultures, in vitro . PNiPAAm–CII-immunized arthritic mouse paws showed massive infiltration of immune cells and extensive damage to cartilage and bone. As determined by immunostaining, most of the CII protein retained its native configuration after injecting it with PNiPAAm in naive mice. Physical adsorption of CII and the high-molecular-weight form of moderately hydrophobic PNiPAAm induced a significant anti-CII antibody response. Similar to CII, mice immunized with PNiPAAm and ovalbumin (PNiPAAm–Ova) induced significant anti-ovalbumin antibody response. Comparable levels of serum IFN-γ, IL-1β and IL-17 were observed in ovalbumin-immunized mice with complete Freund, incomplete Freund (CFA and IFA) or PNiPAAm adjuvants. However, serum IL-4 levels were significantly higher in PNiPAAm–Ova and CFA–Ova groups compared with the IFA–Ova group. Thus, we show for the first time, biocompatible and biodegradable thermo-responsive PNiPAAm can be used as an adjuvant in several immunological applications as well as in better understanding of the autoimmune responses against self-proteins.

scholarly journals Type I and Type II Interferons Inhibit the Translation of Murine Norovirus Proteins

2009 ◽  
Vol 83 (11) ◽  
pp. 5683-5692 ◽  
Author(s):  
Harish Changotra ◽  
Yali Jia ◽  
Tara N. Moore ◽  
Guangliang Liu ◽  
Shannon M. Kahan ◽  
...  
Keyword(s):  
Viral Replication ◽  
Small Animal ◽  
Type I ◽  
Type Ii ◽  
Murine Norovirus ◽  
Human Noroviruses ◽  
Interferon Responses ◽  
Replication Intermediates

ABSTRACT Human noroviruses are responsible for more than 95% of nonbacterial epidemic gastroenteritis worldwide. Both onset and resolution of disease symptoms are rapid, suggesting that components of the innate immune response are critical in norovirus control. While the study of the human noroviruses has been hampered by the lack of small animal and tissue culture systems, our recent discovery of a murine norovirus (MNV) and its in vitro propagation have allowed us to begin addressing norovirus replication strategies and immune responses to norovirus infection. We have previously demonstrated that interferon responses are critical to control MNV-1 infection in vivo and to directly inhibit viral replication in vitro. We now extend these studies to define the molecular basis for interferon-mediated inhibition. Viral replication intermediates were not detected in permissive cells pretreated with type I interferon after either infection or transfection of virion-associated RNA, demonstrating a very early block to virion production that is after virus entry and uncoating. A similar absence of viral replication intermediates was observed in infected primary macrophages and dendritic cells pretreated with type I IFN. This was not due to degradation of incoming genomes in interferon-pretreated cells since similar levels of genomes were present in untreated and pretreated cells through 6 h of infection, and these genomes retained their integrity. Surprisingly, this block to the translation of viral proteins was not dependent on the well-characterized interferon-induced antiviral molecule PKR. Similar results were observed in cells pretreated with type II interferon, except that the inhibition of viral translation was dependent on PKR. Thus, both type I and type II interferon signaling inhibit norovirus translation in permissive myeloid cells, but they display distinct dependence on PKR for this inhibition.

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