Immunotherapy-related cognitive impairment after CAR T cell therapy in mice.

Persistent central nervous system (CNS) immune dysregulation and consequent dysfunction of multiple neural cell types is central to the neurobiological underpinnings of a cognitive impairment syndrome that can occur following traditional cancer therapies or certain infections. Immunotherapies have revolutionized cancer care for many tumor types, but the potential long-term cognitive sequelae are incompletely understood. Here, we demonstrate in mouse models that chimeric antigen receptor (CAR) T cell therapy for both CNS and non-CNS cancers can impair cognitive function and induce a persistent CNS immune response characterized by white matter microglial reactivity and elevated cerebrospinal fluid (CSF) cytokines and chemokines. Consequently, oligodendroglial homeostasis and hippocampal neurogenesis are disrupted. Microglial depletion rescues oligodendroglial deficits and cognitive performance in a behavioral test of attention and short-term memory function. Taken together, these findings illustrate similar mechanisms underlying immunotherapy-related cognitive impairment (IRCI) and cognitive impairment following traditional cancer therapies and other immune challenges.

Alleviating psoriatic skin inflammation through augmentation of Treg cells via CTLA-4 signaling peptide.

Psoriasis is a chronic inflammatory skin disease characterized by hyperplasia of keratinocytes and immune cell infiltration. The IL-17-producing T cells play a key role in psoriasis pathogenesis, while regulatory T (Treg) cells are diminished during psoriatic inflammation. Current psoriasis treatments largely focus on IL-17 and IL-23, however, few studies have explored therapeutic drugs targeting an increase of Treg cells to control immune homeostasis. In this study, we investigated the effects of a cytotoxic T lymphocyte antigen-4 (CTLA-4) signaling peptide (dNP2-ctCTLA-4) in Th17, Tc17, γδ T cells, Treg cells in vitro and a mouse model of psoriasis. Treatment with dNP2-ctCTLA-4 peptide showed a significant reduction of psoriatic skin inflammation with increased Treg cell proportion and reduced IL-17 production by T cells, indicating a potential role in modulating psoriatic skin disease. We compared dNP2-ctCTLA-4 with CTLA-4-Ig and found that only dNP2-ctCTLA-4 ameliorated the psoriasis progression, with increased Treg cells and inhibited IL-17 production from γδ T cells. In vitro experiments using a T cell-antigen presenting cell co-culture system demonstrated the distinct mechanisms of dNP2-ctCTLA-4 compared to CTLA-4-Ig in the induction of Treg cells. These findings highlight the therapeutic potential of dNP2-ctCTLA-4 peptide in psoriasis by augmenting Treg/Teff ratio, offering a new approach to modulating the disease.

Co-opting signalling molecules enables logic-gated control of CAR T cells.

Although chimeric antigen receptor (CAR) T cells have altered the treatment landscape for B cell malignancies, the risk of on-target, off-tumour toxicity has hampered their development for solid tumours because most target antigens are shared with normal cells1,2. Researchers have attempted to apply Boolean-logic gating to CAR T cells to prevent toxicity3-5; however, a truly safe and effective logic-gated CAR has remained elusive6. Here we describe an approach to CAR engineering in which we replace traditional CD3ζ domains with intracellular proximal T cell signalling molecules. We show that certain proximal signalling CARs, such as a ZAP-70 CAR, can activate T cells and eradicate tumours in vivo while bypassing upstream signalling proteins, including CD3ζ. The primary role of ZAP-70 is to phosphorylate LAT and SLP-76, which form a scaffold for signal propagation. We exploited the cooperative role of LAT and SLP-76 to engineer logic-gated intracellular network (LINK) CAR, a rapid and reversible Boolean-logic AND-gated CAR T cell platform that outperforms other systems in both efficacy and prevention of on-target, off-tumour toxicity. LINK CAR will expand the range of molecules that can be targeted with CAR T cells, and will enable these powerful therapeutic agents to be used for solid tumours and diverse diseases such as autoimmunity7 and fibrosis8. In addition, this work shows that the internal signalling machinery of cells can be repurposed into surface receptors, which could open new avenues for cellular engineering.

Fermentative effects by probiotic Lactobacillus brevis B7 on antioxidant and anti-inflammatory properties of hydroponic ginseng.

Soil-cultivation presents environmental limitations and requires considerable labor, space, and water supply. Alternatively, hydroponically-cultured ginseng (HG) was improved its productivity, availability, and functionality. Improvement of bio-functionality by probiotic fermentation also has been studied. Therefore, in this study, HG was fermented using probiotics to enhance antioxidant and anti-inflammatory activities. Soil-cultivated ginseng (SG), 1 and 2-year HG (HG1, HG2) were extracted using 70% ethanol and fermented by Lactobacillus brevis B7. After fermentation, the phenolic and flavonoid contents, and antioxidant and NO scavenging activities were increased, and HG showed higher bioactivities than SG. Particularly, fermented HG2 showed the highest antioxidant and anti-inflammatory activities and significantly decreased the level of inflammatory mediators. Furthermore, fermented HG2 also effectively inhibited NF-κB signaling pathway. These results suggested that fermented HG significantly enhanced functionality compared to SG and non-fermented HG. This suggests that fermented HG is a potentially useful ingredient for developing health-functional foods or pharmaceutical materials.

Levilactobacillus brevis KU15151 Inhibits Staphylococcus aureus Lipoteichoic Acid-Induced Inflammation in RAW 264.7 Macrophages.

Inflammation is a host defense response to harmful agents, such as pathogenic invasion, and is necessary for health. Excessive inflammation may result in the development of inflammatory disorders. Levilactobacillus brevis KU15151 has been reported to exhibit probiotic characteristics and antioxidant activities, but the effect of this strain on inflammatory responses has not been determined. The present study aimed to investigate the anti-inflammatory potential of L. brevis KU15151 in Staphylococcus aureus lipoteichoic acid (aLTA)-induced RAW264.7 macrophages. Treatment with L. brevis KU15151 reduced the production of nitric oxide and prostaglandin E2 by suppressing the expression of inducible nitric oxide synthase and cyclooxygenase-2. Additionally, the production of proinflammatory cytokines including tumor necrosis factor-α, interleukin (IL)-6, and IL-1ß, decreased after treatment with L. brevis KU15151 in aLTA-stimulated RAW 264.7 cells. Furthermore, this strain alleviated the activation of nuclear factor-κB and mitogen-activated protein kinase signaling pathways. Moreover, the generation of reactive oxygen species was downregulated by treatment with L. brevis KU15151. These results demonstrate that L. brevis KU15151 possesses an inhibitory effect against aLTA-mediated inflammation and may be employed as a functional probiotic for preventing inflammatory disorders.

Optimization of an Industrial Medium and Culture Conditions for Probiotic Weissella cibaria JW15 Biomass Using the Plackett-Burman Design and Response Surface Methodology.

The objective of this study was to optimize industrial-grade media for improving the biomass production of Weissella cibaria JW15 (JW15) using a statistical approach. Eleven variables comprising three carbon sources (glucose, fructose, and sucrose), three nitrogen sources (protease peptone, yeast extract, and soy peptone), and five mineral sources (K2HPO4, potassium citrate, L-cysteine phosphate, MgSO4, and MnSO4) were screened by using the Plackett-Burman design. Consequently, glucose, sucrose, and soy peptone were used as significant variables in response surface methodology (RSM). The composition of the optimal medium (OM) was 22.35 g/l glucose, 15.57 g/l sucrose, and 10.05 g/l soy peptone, 2.0 g/l K2HPO4, 5.0 g/l sodium acetate, 0.1 g/l MgSO4·7H2O, 0.05 g/l MnSO4·H2O, and 1.0 g/l Tween 80. The OM significantly improved the biomass production of JW15 over an established commercial medium (MRS). After fermenting OM, the dry cell weight of JW15 was 4.89 g/l, which was comparable to the predicted value (4.77 g/l), and 1.67 times higher than that of the MRS medium (3.02 g/l). Correspondingly, JW15 showed a rapid and increased production of lactic and acetic acid in the OM. To perform a scale-up validation, batch fermentation was executed in a 5-l bioreactor at 37°C with or without a pH control at 6.0 ± 0.1. The biomass production of JW15 significantly improved (1.98 times higher) under the pH control, and the cost of OM was reduced by two-thirds compared to that in the MRS medium. In conclusion, OM may be utilized for mass producing JW15 for industrial use.

Protective Effects of a Novel Lactobacillus brevis Strain with Probiotic Characteristics against Staphylococcus aureus Lipoteichoic Acid-Induced Intestinal Inflammatory Response.

Probiotics can effectively modulate host immune responses and prevent gastrointestinal diseases. The objective of this study was to investigate the probiotic characteristics of Lactobacillus brevis KU15152 isolated from kimchi and its protective potential against intestinal inflammation induced by Staphylococcus aureus lipoteichoic acid (aLTA). L. brevis KU15152 exhibited a high survival rate in artificial gastric and bile environments. Additionally, the adhesion capability of the strain to HT-29 cells was higher than that of L. rhamnosus GG. L. brevis KU15152 did not produce harmful enzymes, such as ß-glucuronidase, indicating that it could be used as a potential probiotic. The anti-inflammatory potential of L. brevis KU15152 was determined in HT-29 cells. Treatment with L. brevis KU15152 suppressed the production of interleukin-8 without inducing significant cytotoxicity. The downregulatory effects of L. brevis KU15152 were involved in the suppression of nuclear factor-kappa B activation mediated by the extracellular signal-regulated kinase and Akt signaling pathways. Collectively, these data suggest that L. brevis KU15152 can be used in developing therapeutic and prophylactic products to manage and treat aLTA-induced intestinal damage.

CPP Applications in Immune Modulation and Disease Therapy.

About 30 years ago, the discovery of CPP improved the therapeutic approach to treat diseases and extended the range of potential targets to intracellular molecules. There are potential drug candidates for FDA approval based on active studies in basic research, preclinical, and clinical trials. Various attempts by CPP application to control the diseases such as allergy, autoimmunity, cancer, and infection demonstrated a strategy to make a new drug pipeline for successful discovery of a biologic drug for immune modulation. However, there are still no CPP-based drug candidates for immune-related diseases in the clinical stage. To control immune responses successfully, not only increasing delivery efficiency of CPPs but also selecting potential target cells and cargoes could be important issues. In particular, as it becomes possible to control intracellular targets, efforts to find various novel potential target are being attempted. In this chapter, we focused on CPP-based approaches to treat diseases through modulation of immune responses and discussed for perspectives on future direction of the research for successful application of CPP technology to immune modulation and disease therapy in clinical trial.

The Cysteine-Containing Cell-Penetrating Peptide AP Enables Efficient Macromolecule Delivery to T Cells and Controls Autoimmune Encephalomyelitis.

T cells are key immune cells involved in the pathogenesis of several diseases, rendering them important therapeutic targets. Although drug delivery to T cells is the subject of continuous research, it remains challenging to deliver drugs to primary T cells. Here, we used a peptide-based drug delivery system, AP, which was previously developed as a transdermal delivery peptide, to modulate T cell function. We first identified that AP-conjugated enhanced green fluorescent protein (EGFP) was efficiently delivered to non-phagocytic human T cells. We also confirmed that a nine-amino acid sequence with one cysteine residue was the optimal sequence for protein delivery to T cells. Next, we identified the biodistribution of AP-dTomato protein in vivo after systemic administration, and transduced it to various tissues, such as the spleen, liver, intestines, and even to the brain across the blood-brain barrier. Next, to confirm AP-based T cell regulation, we synthesized the AP-conjugated cytoplasmic domain of CTLA-4, AP-ctCTLA-4 peptide. AP-ctCTLA-4 reduced IL-17A expression under Th17 differentiation conditions in vitro and ameliorated experimental autoimmune encephalomyelitis, with decreased numbers of pathogenic IL-17A+GM-CSF+ CD4 T cells. These results collectively suggest the AP peptide can be used for the successful intracellular regulation of T cell function, especially in the CNS.

Chrysanthemum indicum suppresses adipogenesis by inhibiting mitotic clonal expansion in 3T3-L1 preadipocytes.

Herbs have been of interest to treat diseases, including obesity, owing to their various bioactive constituents that exhibit therapeutic and prophylactic properties. The present study examined the anti-adipogenic effects and mechanisms of Chrysanthemum indicum aqueous extract (CAE) in 3T3-L1 preadipocytes. CAE comprises 1,3-dicaffeoylquinic acid, chlorogenic acid, kaempferol-3-O-glucoside, caffeic acid, and apigenin, which were corresponded with previous reports. CAE inhibited the accumulation of lipid droplets and significantly alleviated the expression of lipogenesis- and adipogenesis-associated biomarkers. Treatment with CAE inhibited the mitotic clonal expansion (MCE), corroborated by cell cycle arrest at the G0 /G1 phase, and mitigated the expression of cell cycle progression-associated proteins and in addition to phosphorylation of MCE-promoting transcription factors. Moreover, CAE downregulated the activation of Akt and extracellular signal-regulated kinase 1/2 signaling pathways. In summary, CAE facilitates adipogenic inhibition during the early phase of differentiation, especially MCE, and its phenolic compounds can contribute to its anti-obesogenic properties. PRACTICAL APPLICATIONS: Chrysanthemum indicum has been mainly used as traditional herbal tea and drinks. Chrysanthemum indicum aqueous extract (CAE) inhibits adipogenesis by suppressing mitotic clonal expansion during the early phase of differentiation in 3T3-L1 preadipocytes. 1,3-Dicaffeoylquinic acid, chlorogenic acid, kaempferol-3-O-glucoside, caffeic acid, and apigenin were detected in CAE. Based on these findings, CAE can be used as nutraceutical agents for prevention and treatment of obesity.

In Vivo Induction of Regulatory T Cells Via CTLA-4 Signaling Peptide to Control Autoimmune Encephalomyelitis and Prevent Disease Relapse.

Regulatory T cells play a key role in immune tolerance to self-antigens, thereby preventing autoimmune diseases. However, no drugs targeting Treg cells have been approved for clinical trials yet. Here, a chimeric peptide is generated by conjugation of the cytoplasmic domain of CTLA-4 (ctCTLA-4) with dNP2 for intracellular delivery, dNP2-ctCTLA-4, and evaluated Foxp3 expression during Th0, Th1, Treg, and Th17 differentiation dependent on TGF-ß. The lysine motif of ctCTLA-4, not tyrosine motif, is required for Foxp3 expression for Treg induction and amelioration of experimental autoimmune encephalomyelitis (EAE). Transcriptome analysis reveals that dNP2-ctCTLA-4-treated T cells express Treg transcriptomic patterns with properties of suppressive functions. In addition, the molecular interaction between the lysine motif of ctCTLA-4 and PKC-η is critical for Foxp3 expression. Although both CTLA-4-Ig and dNP2-ctCTLA-4 treatment in vivo ameliorated EAE progression, only dNP2-ctCTLA-4 requires Treg cells for inhibition of disease progression and prevention of relapse. Furthermore, the CTLA-4 signaling peptide is able to induce human Tregs in vitro and in vivo as well as from peripheral blood mononuclear cells (PBMCs) of multiple sclerosis patients. These results collectively suggest that the chimeric CTLA-4 signaling peptide can be used for successful induction of regulatory T cells in vivo to control autoimmune diseases, such as multiple sclerosis.

Induction of the IL-1RII decoy receptor by NFAT/FOXP3 blocks IL-1ß-dependent response of Th17 cells.

Derived from a common precursor cell, the balance between Th17 and Treg cells must be maintained within immune system to prevent autoimmune diseases. IL-1ß-mediated IL-1 receptor (IL-1R) signaling is essential for Th17-cell biology. Fine-tuning of IL-1R signaling is controlled by two receptors, IL-1RI and IL-RII, IL-1R accessory protein, and IL-1R antagonist. We demonstrate that the decoy receptor, IL-1RII, is important for regulating IL-17 responses in TCR-stimulated CD4+ T cells expressing functional IL-1RI via limiting IL-1ß responsiveness. IL-1RII expression is regulated by NFAT via its interaction with Foxp3. The NFAT/FOXP3 complex binds to the IL-1RII promoter and is critical for its transcription. Additionally, IL-1RII expression is dysregulated in CD4+ T cells from patients with rheumatoid arthritis. Thus, differential expression of IL-1Rs on activated CD4+ T cells defines unique immunological features and a novel molecular mechanism underlies IL-1RII expression. These findings shed light on the modulatory effects of IL-1RII on Th17 responses.

Inula britannica Inhibits Adipogenesis of 3T3-L1 Preadipocytes via Modulation of Mitotic Clonal Expansion Involving ERK 1/2 and Akt Signaling Pathways.

The flower of Inula britannica contains various phenolic compounds with prophylactic properties. This study aimed to determine the anti-adipogenic effect of an I. britannica flower aqueous extract (IAE) and its underlying mechanisms in the 3T3-L1 preadipocytes and to identify the phenolic compounds in the extract. Treatment with IAE inhibited the adipogenesis by showing a dose-dependent suppressed intracellular lipid accumulation and mitigated expression levels of lipogenesis- and adipogenesis-associated biomarkers including transcription factors. IAE exerted an anti-adipogenic effect through the modulation of the early phases of adipogenesis including mitotic clonal expansion (MCE). Treatment with IAE inhibited MCE by arresting the cell cycle at the G0/G1 phase and suppressing the activation of MCE-related transcription factors. Furthermore, IAE inhibited adipogenesis by regulating the extracellular signal-regulated kinase 1/2 and Akt signaling pathways. Protocatechuic acid, chlorogenic acid, kaempferol-3-O-glucoside, and 6-methoxyluteolin, which are reported to exhibit anti-adipogenic properties, were detected in IAE. Therefore, modulation of early phases of adipogenesis, especially MCE, is a key mechanism underlying the anti-adipogenic activity of IAE. In summary, the anti-obesity effects of IAE can be attributed to its phenolic compounds, and hence, IAE can be used for the development of anti-obesity products.

Cell-Penetrating Function of the Poly(ADP-Ribose) (PAR)-Binding Motif Derived from the PAR-Dependent E3 Ubiquitin Ligase Iduna.

Iduna is a poly(ADP-ribose) (PAR)-dependent E3 ubiquitin ligase that regulates cellular responses such as proteasomal degradation and DNA repair upon interaction with its substrate. We identified a highly cationic region within the PAR-binding motif of Iduna; the region was similar among various species and showed amino acid sequence similarity with that of known cell-penetrating peptides (CPPs). We hypothesized that this Iduna-derived cationic sequence-rich peptide (Iduna) could penetrate the cell membrane and deliver macromolecules into cells. To test this hypothesis, we generated recombinant Iduna-conjugated enhanced green fluorescent protein (Iduna-EGFP) and its tandem-repeat form (d-Iduna-EGFP). Both Iduna-EGFP and d-Iduna-EGFP efficiently penetrated Jurkat cells, with the fluorescence signals increasing dose- and time-dependently. Tandem-repeats of Iduna and other CPPs enhanced intracellular protein delivery efficiency. The delivery mechanism involves lipid-raft-mediated endocytosis following heparan sulfate interaction; d-Iduna-EGFP was localized in the nucleus as well as the cytoplasm, and its residence time was much longer than that of other controls such as TAT and Hph-1. Moreover, following intravenous administration to C57/BL6 mice, d-Iduna-EGFP was efficiently taken up by various tissues, including the liver, spleen, and intestine suggesting that the cell-penetrating function of the human Iduna-derived peptide can be utilized for experimental and therapeutic delivery of macromolecules.

Protein tyrosine phosphatase conjugated with a novel transdermal delivery peptide, astrotactin 1-derived peptide recombinant protein tyrosine phosphatase (AP-rPTP), alleviates both atopic dermatitis-like and psoriasis-like dermatitis.

BACKGROUND: Atopic dermatitis (AD) and psoriasis are the 2 most common chronic inflammatory skin diseases. There is an unmet medical need to overcome limitations for transcutaneous drug development posed by the skin barrier. OBJECTIVE: We aimed to identify a novel transdermal delivery peptide and to develop a transcutaneously applicable immunomodulatory protein for treating AD and psoriasis. METHODS: We identified and generated reporter proteins conjugated to astrotactin 1-derived peptide (AP), a novel transdermal delivery peptide of human origin, and analyzed the intracellular delivery efficiency of these proteins in mouse and human skin cells and tissues using multiphoton confocal microscopy. We also generated a recombinant therapeutic protein, AP-recombinant protein tyrosine phosphatase (rPTP), consisting of the phosphatase domain of the T-cell protein tyrosine phosphatase conjugated to AP. The immunomodulatory function of AP-rPTP was confirmed in splenocytes on cytokine stimulation and T-cell receptor stimulation. Finally, we confirmed the in vivo efficacy of AP-rPTP transdermal delivery in patients with oxazolone-induced contact hypersensitivity, ovalbumin-induced AD-like, and imiquimod-induced psoriasis-like skin inflammation models. RESULTS: AP-conjugated reporter proteins exhibited significant intracellular transduction efficacy in keratinocytes, fibroblasts, and immune cells. In addition, transcutaneous administration of AP-dTomato resulted in significant localization into the dermis and epidermis in both mouse and human skin. AP-rPTP inhibited phosphorylated signal transducer and activator of transcription (STAT) 1, STAT3, and STAT6 in splenocytes and also regulated T-cell activation and proliferation. Transcutaneous administration of AP-rPTP through the paper-patch technique significantly ameliorated skin tissue thickening, inflammation, and cytokine expression in both AD-like and psoriasis-like dermatitis models. CONCLUSION: We identified a 9-amino-acid novel transdermal delivery peptide, AP, and demonstrated its feasibility for transcutaneous biologic drug development. Moreover, AP-rPTP is a novel immunomodulatory drug candidate for human dermatitis.

dNP2 is a blood-brain barrier-permeable peptide enabling ctCTLA-4 protein delivery to ameliorate experimental autoimmune encephalomyelitis.

Central nervous system (CNS)-infiltrating effector T cells play critical roles in the development and progression of multiple sclerosis (MS). However, current drugs for MS are very limited due to the difficulty of delivering drugs into the CNS. Here we identify a cell-permeable peptide, dNP2, which efficiently delivers proteins into mouse and human T cells, as well as various tissues. Moreover, it enters the brain tissue and resident cells through blood vessels by penetrating the tightly organized blood-brain barrier. The dNP2-conjugated cytoplasmic domain of cytotoxic T-lymphocyte antigen 4 (dNP2-ctCTLA-4) negatively regulates activated T cells and shows inhibitory effects on experimental autoimmune encephalomyelitis in both preventive and therapeutic mouse models, resulting in the reduction of demyelination and CNS-infiltrating T helper 1 and T helper 17 cells. Thus, this study demonstrates that dNP2 is a blood-brain barrier-permeable peptide and dNP2-ctCTLA-4 could be an effective agent for treating CNS inflammatory diseases such as MS.

Comparison between open plating versus minimally invasive plate osteosynthesis for acute displaced clavicular shaft fractures.

BACKGROUND: Current literatures describe good clinical outcomes of acute displaced fracture of clavicle treated with minimally invasive plate osteosynthesis (MIPO). But, there are little comparative data of the outcomes between open plating and MIPO techniques. We compared the outcomes of open plating and MIPO for treatment of acute displaced clavicular shaft fractures. MATERIALS AND METHODS: The author performed a retrospective review on a consecutive series of patients with clavicular shaft fracture who underwent open plating or MIPO. Fourteen patients were treated with open plating with interfragmentary screw fixation, and 19 were treated with the MIPO technique without exposing a fracture site itself. A superior plating method was applied to both groups. Patient demographics, clinical outcomes using Constant score and University of California Los Angeles (UCLA) shoulder score, operation time, union rate, complications, and radiographic evaluation were evaluated. RESULTS: There were no statistically significant differences in the demographic data, including patient's variables (age, gender, involved side, smoking, alcohol, and diabetic status) and fracture characteristics (trauma mechanism, distribution of fracture type, presence of polytrauma, and time from trauma to surgery) between the two groups. Mean operation time was 87.5 min in open plating and 77.2 min in MIPO (p=0.129). The mean time to union was 15.7 weeks in patients who underwent open plating and 16.8 weeks in patients who underwent MIPO (p=0.427). Although there was no significant difference, nonunion developed 1 case in MIPO while none was in open plating. Four patients in open plating had skin numbness (none in MIPO, p=0.024). There was no significant difference in the Constant score and UCLA score of the two surgical methods. CONCLUSION: This study showed that both open plating with interfragmentary screw fixation (Open plating) and minimally invasive plate osteosynthesis (MIPO) are equally effective and safe treatment methods for acute displaced clavicle shaft fracture.

Arthroscopic Partial Repair of Irreparable Rotator Cuff Tears: Preoperative Factors Associated With Outcome Deterioration Over 2 Years.

BACKGROUND: Arthroscopic partial repair is a treatment option in irreparable large-to-massive rotator cuff tears without arthritic changes. However, there are indications that arthroscopic partial repair does not yield satisfactory outcomes. PURPOSE: To report the clinical and radiographic results of arthroscopic partial repairs in patients with irreparable large-to-massive cuff tears. In addition, an analysis was performed regarding preoperative factors that may influence patient outcomes and patient-rated satisfaction over time. STUDY DESIGN: Case series; Level of evidence, 4. METHODS: From 2005 to 2011, a total of 31 patients who underwent arthroscopic partial repair for irreparable large-to-massive cuff tears were retrospectively evaluated. Partial repair was defined as posterior cuff tissue repair with or without subscapularis tendon repair to restore the transverse force couple of the cuff. Pain visual analog scale (PVAS), questionnaire results (American Shoulder and Elbow Surgeons [ASES] and Simple Shoulder Test [SST]), and radiographic changes (acromiohumeral distance and degenerative change) were assessed preoperatively, at first follow-up (roughly 1 year postoperatively), and at final follow-up (>2 years postoperatively). Patients rated their satisfaction level at each postoperative follow-up as well. Preoperative factors that might influence outcomes, such as patient demographics, tear size, and fatty infiltration, were investigated. RESULTS: The preoperative, first follow-up, and final follow-up results for mean PVAS (5.13, 2.13, and 3.16, respectively) and questionnaires (ASES: 41.97, 76.37, and 73.78; SST: 3.61, 6.33, and 6.07, respectively) improved significantly (all P < .05). Radiographic evaluation showed no difference compared with preoperative status. Nevertheless, patient-rated satisfaction at final evaluation was inferior: 16 good responses ("very satisfied" and "satisfied") and 15 poor responses ("rather the same" and "dissatisfied"). Despite initial improvements in both groups (P < .05), patients with poor satisfaction demonstrated statistically significant deterioration in mean PVAS (from 2.07 to 4.67), questionnaire scores (ASES: from 74.56 to 59.80; SST: from 5.11 to 3.81), and acromiohumeral distance (from 7.19 to 5.06 mm) between the first and final follow-up (all P < .05). Patients with good satisfaction showed no significant difference or they improved (P > .05) from the first to the final follow-up. Among preoperative factors, fatty infiltration of the teres minor was identified as the only statistically significant factor affecting patient-rated satisfaction (P = .007). CONCLUSION: This study showed that arthroscopic partial repair may produce initial improvement in selected outcomes at 2-year follow-up. However, about half of the patients in the study were not satisfied with their outcomes, which had deteriorated over time. Preoperative fatty infiltration of the teres minor was the only factor that correlated with worse final outcomes and poor satisfaction after arthroscopic partial repair.

Cell membrane penetrating function of the nuclear localization sequence in human cytokine IL-1α.

Cytokines are released from the cell, bind to their receptors, and affect cellular responses. The precursor form of interleukin 1 alpha (pIL-1α) has a nuclear localization sequence (NLS) that causes it to be localized to the nucleus and regulate specific gene expression. The amino acids of the NLS are basic amino acid-rich sequences, as is the cell penetrating peptide (CPP), which has been widely studied as a way to deliver macromolecules into cells. Here, we hypothesized that the NLS in pIL-1α (pIL-1αNLS) can penetrate the cell membrane and it could deliver macromolecules such as protein in vivo. We characterized cell membrane penetration ability of pIL-1αNLS or its tandem repeated form (2pIL-1αNLS) to enhance its intracellular delivery efficiency. 2pIL-1αNLS showed comparable protein delivery efficiency to TAT-CPP and it mediates endocytosis following heparan sulfate interaction. 2pIL-1αNLS conjugated enhanced green fluorescence protein was localized to the nucleus and the cytoplasm. Intra-peritoneal administration of 2pIL-1αNLS conjugated dTomato protein showed remarkable in vivo intracellular delivery efficiency in various tissues including spleen, liver, and intestine in mice. Moreover, cytotoxicity of 2pIL-1αNLS was not observed even at 100 µM. Our results demonstrate cell membrane-penetrating function of NLS in pIL-1α, which can be used as a safe therapeutic macromolecular delivery peptide.

PPARγ negatively regulates T cell activation to prevent follicular helper T cells and germinal center formation.

Peroxisome proliferator-activated receptor gamma (PPARγ) is a transcription factor that regulates lipid and glucose metabolism. Although studies of PPARγ ligands have demonstrated its regulatory functions in inflammation and adaptive immunity, its intrinsic role in T cells and autoimmunity has yet to be fully elucidated. Here we used CD4-PPARγKO mice to investigate PPARγ-deficient T cells, which were hyper-reactive to produce higher levels of cytokines and exhibited greater proliferation than wild type T cells with increased ERK and AKT phosphorylation. Diminished expression of IκBα, Sirt1, and Foxo1, which are inhibitors of NF-κB, was observed in PPARγ-deficient T cells that were prone to produce all the signature cytokines under Th1, Th2, Th17, and Th9 skewing condition. Interestingly, 1-year-old CD4-PPARγKO mice spontaneously developed moderate autoimmune phenotype by increased activated T cells, follicular helper T cells (TFH cells) and germinal center B cells with glomerular inflammation and enhanced autoantibody production. Sheep red blood cell immunization more induced TFH cells and germinal centers in CD4-PPARγKO mice and the T cells showed increased of Bcl-6 and IL-21 expression suggesting its regulatory role in germinal center reaction. Collectively, these results suggest that PPARγ has a regulatory role for TFH cells and germinal center reaction to prevent autoimmunity.