Supercharged coiled-coil protein with N-terminal decahistidine tag boosts siRNA complexation and delivery efficiency of a lipoproteoplex.

Short interfering RNA (siRNA) therapeutics have soared in popularity due to their highly selective and potent targeting of faulty genes, providing a non-palliative approach to address diseases. Despite their potential, effective transfection of siRNA into cells requires the assistance of an accompanying vector. Vectors constructed from non-viral materials, while offering safer and non-cytotoxic profiles, often grapple with lackluster loading and delivery efficiencies, necessitating substantial milligram quantities of expensive siRNA to confer the desired downstream effects. We detail the recombinant synthesis of a diverse series of coiled-coil supercharged protein (CSP) biomaterials systematically designed to investigate the impact of two arginine point mutations (Q39R and N61R) and decahistidine tags on liposomal siRNA delivery. The most efficacious variant, N8, exhibits a twofold increase in its affinity to siRNA and achieves a twofold enhancement in transfection activity with minimal cytotoxicity in vitro. Subsequent analysis unveils the destabilizing effect of the Q39R and N61R supercharging mutations and the incorporation of C-terminal decahistidine tags on α-helical secondary structure. Cross-correlational regression analyses reveal that the amount of helical character in these mutants is key in N8's enhanced siRNA complexation and downstream delivery efficiency.

Osteoclasts and Probiotics Mediate Significant Expansion, Functional Activation and Supercharging in NK, γδ T, and CD3+ T Cells: Use in Cancer Immunotherapy.

Our previous studies have introduced osteoclasts (OCs) as major activators of NK cells. It was found that OCs exhibit the capabilities of inducing cell expansion as well as increasing the cytotoxic activity of NK cells by granule release and increasing the secretion of TNF-α and TRAIL, leading to increased lysis of tumors in short-term as well as long-term periods, respectively. OC- induced expanded NK cells were named supercharged NK cells (sNK) due to their significantly high functional activity as well as their significantly higher cell expansion rate. It is, however, unclear whether the OC-mediated effect in NK cells is specific or whether other cytotoxic immune cells can also be expanded and activated by OCs. We chose to focus on γδ T cells and pan T cells, which also include CD8+ T cells. In this paper, we report that OCs are capable of expanding and functionally activating both γδ T cells and pan T cells. Expanded γδ T and pan T cells were capable of secreting high levels of INF-γ, albeit with different dynamics to those of NK cells, and, moreover, they are unable to kill NK-specific targets. Since we used humanized-BLT (hu-BLT) mice as a model of human disease, we next determined whether NK and T cell activation through OCs is also evident in cells obtained from hu-BLT mice. Similar to humans, OCs were capable of increasing the cell expansion and secretion of IFN-γ in the culture of either NK or T cells from hu-BLT mice, providing yet further evidence that these mice are appropriate models to study human disease. Therefore, these studies indicated that CD3+ T or γδ T cells can proliferate and be supercharged by OCs similar to the NK cells; thus, they can be used individually or in combination in the cell therapy of cancers.

Supercharging of the ulnar nerve: clinical and neurophysiological assessment at 2 years for nine proximal injuries.

LEVEL OF EVIDENCE: II.

Supercharged End-to-Side Nerve Transfer for Ulnar Neuropathy: Redefining Nomenclature and Recommendations for Standardisation of Surgical Technique Description.

Supercharged end-to-side (SETS) nerve transfers have been described as a treatment option for ulnar neuropathy, however, there is inconsistency in the nomenclature used to describe the microsurgical technique. The purpose of this article is to systematically review the available literature on the SETS nerve transfer technique and to provide an overview of the technical variations to facilitate standardisation of surgical method. A literature review was performed through PubMed, MEDLINE, and Ovid databases according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. Studies that reported surgical technique of anterior interosseous nerve (AIN) to ulnar nerve SETS transfer were included. Studies were excluded when not referencing SETS/reverse end-to-side (RETS) nerve transfers, studies referencing nerve transfers other than AIN to motor fascicle bundle of the ulnar nerve (MUN), animal studies, and studies not reporting technique. Of the 168 studies found, 14 articles were included. In five articles, distal visualisation of the MUN in Guyon's canal was specifically cited. In the four studies that commented on donor preparation, sharp neurectomy proximal to the AIN branching point was undertaken. Recipient preparation was commented on in seven of the included studies. Two studies referred to an epineurial window only while five specifically recommended a perineurial window. Coaptation site was specified in four studies and all studies used sutures for coaptation, with four articles stipulating that 9-0 nylon was used. Additionally, fibrin glue was used in conjunction with suture technique in four studies. Consistency in nomenclature used to describe SETS microsurgical technique is needed before case series measuring outcome can be reliably interpreted. This review allowed for the development of suggestions for standardisation of nomenclature and minimal reporting requirements when describing SETS technique. Standardisation of technique will allow for reproducibility and facilitate future evaluations of outcome in prospective randomised control trials.

Supercharged Fluorescent Protein-Apoferritin Cocrystals for Lighting Applications.

The application of fluorescent proteins (FPs) in optoelectronics is hindered by the need for effective protocols to stabilize them under device preparation and operational conditions. Factors such as high temperatures, irradiation, and organic solvent exposure contribute to the denaturation of FPs, resulting in a low device performance. Herein, we focus on addressing the photoinduced heat generation associated with FP motion and rapid heat transfer. This leads to device temperatures of approximately 65 °C, causing FP-denaturation and a subsequent loss of device functionality. We present a FP stabilization strategy involving the integration of electrostatically self-assembled FP-apoferritin cocrystals within a silicone-based color down-converting filter. Three key achievements characterize this approach: (i) an engineering strategy to design positively supercharged FPs (+22) without compromising photoluminescence and thermal stability compared to their native form, (ii) a carefully developed crystallization protocol resulting in highly emissive cocrystals that retain the essential photoluminescence features of the FPs, and (iii) a strong reduction of the device's working temperature to 40 °C, leading to a 40-fold increase in Bio-HLEDs stability compared to reference devices.

Sequential therapy with supercharged NK cells with either chemotherapy drug cisplatin or anti-PD-1 antibody decreases the tumor size and significantly enhances the NK function in Hu-BLT mice.

Introduction and methods: In this study we report that sequential treatment of supercharged NK (sNK) cells with either chemotherapeutic drugs or check-point inhibitors eliminate both poorly differentiated and well differentiated tumors in-vivo in humanized-BLT mice. Background and results: sNK cells were found to be a unique population of activated NK cells with genetic, proteomic, and functional attributes that are very different from primary untreated or IL-2 treated NK cells. Furthermore, NK-supernatant differentiated or well-differentiated oral or pancreatic tumor cell lines are not susceptible to IL-2 activated primary NK cell-mediated cytotoxicity; however, they are greatly killed by the CDDP and paclitaxel in in-vitro assays. Injection of one dose of sNK cells at 1 million cells per mouse to aggressive CSC-like/poorly differentiated oral tumor bearing mice, followed by an injection of CDDP, inhibited tumor weight and growth, and increased IFN-γ secretion as well as NK cell-mediated cytotoxicity substantially in bone marrow, spleen and peripheral blood derived immune cells. Similarly, the use of check point inhibitor anti-PD-1 antibody increased IFN-γ secretion and NK cell-mediated cytotoxicity, and decreased the tumor burden in-vivo, and tumor growth of resected minimal residual tumors from hu-BLT mice when used sequentially with sNK cells. The addition of anti-PDL1 antibody to poorly differentiated MP2, NK-differentiated MP2 or well-differentiated PL-12 pancreatic tumors had different effects on tumor cells depending on the differentiation status of the tumor cells, since differentiated tumors expressed PD-L1 and were susceptible to NK cell mediated ADCC, whereas poorly differentiated OSCSCs or MP2 did not express PD-L1 and were killed directly by the NK cells. Conclusions: Therefore, the ability to target combinatorially clones of tumors with NK cells and chemotherapeutic drugs or NK cells with checkpoint inhibitors at different stages of tumor differentiation may be crucial for successful eradication and cure of cancer. Furthermore, the success of check point inhibitor PD-L1 may relate to the levels of expression on tumor cells.

Pharyngeaoesophageal Reconstruction.

Pharyngoesophageal reconstruction is one of the most challenging reconstructive dilemmas that demands extensive planning, meticulous surgical execution, and timely management of postoperative complications. The main goals of reconstruction are to protect critical blood vessels of the neck, to provide alimentary continuity, and to restore functions such as speech and swallowing. With the evolution of techniques, fasciocutaneous flaps have become the gold standard for most defects in this region. Major complications include anastomotic strictures and fistulae, but most patients can tolerate an oral diet and achieve fluent speech after rehabilitation with a tracheoesophageal puncture.

Supercharged Protein Nanosheets for Cell Expansion on Bioemulsions.

Cell culture at liquid-liquid interfaces, for example, at the surface of oil microdroplets, is an attractive strategy to scale up adherent cell manufacturing while replacing the use of microplastics. Such a process requires the adhesion of cells at interfaces stabilized and reinforced by protein nanosheets displaying not only high elasticity but also presenting cell adhesive ligands able to bind integrin receptors. In this report, supercharged albumins are found to form strong elastic protein nanosheets when co-assembling with the co-surfactant pentafluorobenzoyl chloride (PFBC) and mediate extracellular matrix (ECM) protein adsorption and cell adhesion. The interfacial mechanical properties and elasticity of supercharged nanosheets are characterized by interfacial rheology, and behaviors are compared to those of native bovine serum albumin, human serum albumin, and α-lactalbumin. The impact of PFBC on such assembly is investigated. ECM protein adsorption to resulting supercharged nanosheets is then quantified via surface plasmon resonance and fluorescence microscopy, demonstrating that the dual role supercharged albumins are proposed to play as scaffold protein structuring liquid-liquid interfaces and substrates for the capture of ECM molecules. Finally, the adhesion and proliferation of primary human epidermal stem cells are investigated, at pinned droplets, as well as on bioemulsions stabilized by corresponding supercharged nanosheets. This study demonstrates the potential of supercharged proteins for the engineering of biointerfaces for stem cell manufacturing and draws structure-property relationships that will guide further engineering of associated systems.

Beyond the Cubital Tunnel: Use of Adjunctive Procedures in the Management of Cubital Tunnel Syndrome.

BACKGROUND: Our management of cubital tunnel syndrome has expanded to involve multiple adjunctive procedures, including supercharged end-to-side anterior interosseous to ulnar nerve transfer, cross-palm nerve grafts from the median to ulnar nerve, and profundus tenodesis. We also perform intraoperative brief electrical stimulation in patients with severe disease. The aims of this study were to evaluate the impact of adjunctive procedures and electrical stimulation on patient outcomes. METHODS: We performed a retrospective review of 136 patients with cubital tunnel syndrome who underwent operative management from 2013 to 2018. A total of 38 patients underwent adjunctive procedure(s), and 33 received electrical stimulation. A historical cohort of patients who underwent cubital tunnel surgery from 2009 to 2011 (n = 87) was used to evaluate the impact of adjunctive procedures. Study outcomes were postoperative improvements in Disabilities of the Arm, Shoulder, and Hand (DASH) questionnaire scores, pinch strength, and patient-reported pain and quality of life. RESULTS: In propensity score-matched samples, patients who underwent adjunctive procedures had an 11.3-point greater improvement in DASH scores than their matched controls (P = .0342). In addition, patients who received electrical stimulation had significantly improved DASH scores relative to baseline (11.7-point improvement, P < .0001), whereas their control group did not. However, when compared between treatment arms, there were no significant differences for any study outcome. CONCLUSIONS: Patients who underwent adjunctive procedures experienced greater improvement in postoperative DASH scores than their matched pairs. Additional studies are needed to evaluate the effects of brief electrical stimulation in compression neuropathy.

Papel de la yeyunoplastia en la reconstrucción esofágica compleja / Role of jejunoplasty in complex esophageal reconstruction

Introducción: La reconstrucción esofágica es un proceso quirúrgico técnicamente muy complejo, gravado por una importante morbilidad. Clásicamente se han utilizado la gastroplastia y la coloplastia, aunque la yeyunoplastia ya fue descrita por Roux en 1907. Parece demostrado que la plastia de yeyuno libre es una muy buena opción en el tratamiento de la enfermedad del esófago cervical, pero no está tan claro el papel de la yeyunoplastia supercharged en la reconstrucción del esófago torácico. El objetivo de este estudio es el análisis de las reconstrucciones esofágicas realizadas en nuestra unidad y que precisaron de un injerto de yeyuno. Métodos: Estudio retrospectivo de las reconstrucciones esofágicas realizadas con yeyunoplastias en nuestra unidad entre enero de 2011 y diciembre de 2019. Se analizan datos epidemiológicos, indicaciones, técnica quirúrgica y morbimortalidad. Resultados: Se realizaron 67 procedimientos quirúrgicos de reconstrucción esofágica compleja de los que 10 fueron yeyunoplastias: 5 yeyunos libres en esófago cervical y 5 supercharged en esófago torácico con abordaje transesternal. La morbilidad, mortalidad, estancia media y tiempo de retirada de la alimentación enteral fueron menores en los yeyunos libres que en los supercharged. Conclusiones: En nuestro grupo la yeyunoplastia supercharged es la última opción para la reconstrucción del esófago torácico; el acceso por esternotomía media nos permite un excelente abordaje del mediastino anterior y los vasos mamarios internos. El yeyuno libre sería la primera elección —con indemnidad del resto de esófago— en la reconstrucción del esófago cervical. (AU)

Introduction: Esophageal reconstruction is a very complex surgical procedure, burdened by significant morbidity. Gastroplasty and coloplasty have classically been used. Free jejunal plasty has shown to be a very good option in the treatment of cervical esophagus pathology, but the role of supercharged jejunoplasty in thoracic esophagus reconstruction is still controversial. Methods: A retrospective study of esophageal reconstructions with jejunoplasties performed in our unit between January 2011 and December 2019. Epidemiological data, indications, surgical technique, and morbidity and mortality were analyzed. Results: 67 procedures of esophageal reconstruction were performed, 10 of which were jejunoplasties: 5 free jejunums and 5 supercharged. Morbidity, mortality, mean stay and withdrawal time from enteral feeding were lower in free than in supercharged jejunums. Conclusions: Supercharged jejunoplasty was the last option for reconstruction of the thoracic esophagus. Median sternotomy access provides an excellent approach to the anterior mediastinum and the internal mammary vessels. The free jejunum would be the first choice, with the indemnity of the rest of the esophagus, in the reconstruction of the cervical esophagus. (AU)

Predicting transport of intra-articularly injected growth factor fusion proteins into human knee joint cartilage.

There are no drugs or treatment methods known to prevent the development of post-traumatic osteoarthritis (PTOA), a type of osteoarthritis (OA) that is triggered by traumatic joint injuries and accounts for ∼12% of the nearly 600 million OA cases worldwide. Lack of effective drug delivery techniques remains a major challenge in developing clinically effective treatments, but cationic delivery carriers can help overcome this challenge. Scaling up treatments that are effective in in vitro models to achieve success in preclinical in vivo models and clinical trials is also a challenging problem in the field. Here we use a cationic green fluorescent protein (GFP) as a carrier to deliver Insulin-Like Growth Factor 1 (IGF-1), a drug considered as a potential therapeutic for PTOA. GFP-IGF-1 conjugates were first synthesized as fusion proteins with different polypeptide linkers, and their transport properties were characterized in human cartilage explants. In vitro experimental data were used to develop a predictive mathematical transport model that was validated using an independent in vitro experimental data set. The model was used to predict the transport of these fusion proteins upon intra-articular injection into human knee joints. The predictions included results for the rate and extent of fusion protein penetration into cartilage, and the maximum levels of fusion proteins that would escape into systemic circulation through the joint capsule. Together, our transport measurements and model set the stage for translation of such explant culture studies to in vivo preclinical studies and potentially clinical application. STATEMENT OF SIGNIFICANCE: The lack of blood supply in cartilage and rapid clearance of drugs injected into human knees presents a major challenge in developing clinically effective treatments for osteoarthritis. Cationic delivery carriers can target negatively charged cartilage and help overcome this problem. Scaling up treatments that are effective in vitro to achieve success in vivo is also challenging. Here, we use a cationic green fluorescent protein (GFP) to deliver Insulin-Like Growth Factor-1 (IGF-1) into cartilage. Experiments measuring transport of GFP-IGF-1 fusion proteins in human cartilage explants were used to develop and validate a mathematical model to predict fusion protein transport upon injection into human knee joints. This work translates such explant culture studies to in vivo preclinical studies and potentially clinical application.

Supercharged Jejunal Interposition.

Complex esophageal reconstruction represents a high risk and challenging procedure. A dedicated pathway with multispecialty teams can facilitate a systematic checklist approach to perioperative management and evaluation of long-term outcomes. Refinements in the operative technique for supercharged pedicled jejunum (SPJ) for long segment interposition in esophageal reconstruction are reviewed in this article. Medical and surgical complications among this complex niche group of patients are significant and require care in specialist centers with a focused team. Patient-reported outcomes (PROs) in long-segment SPJ interposition are recognized to provide additional monitoring of surgical outcomes and may help guide interventions for subsequent symptom control.

Role of jejunoplasty in complex esophageal reconstruction.

INTRODUCTION: Esophageal reconstruction is a very complex surgical procedure, burdened by significant morbidity. Gastroplasty and coloplasty have classically been used. Free jejunal plasty has shown to be a very good option in the treatment of cervical esophagus pathology, but the role of supercharged jejunoplasty in thoracic esophagus reconstruction is still controversial. METHODS: A retrospective study of esophageal reconstructions with jejunoplasties performed in our unit between January 2011 and December 2019. Epidemiological data, indications, surgical technique, and morbidity and mortality were analyzed. RESULTS: 67 procedures of esophageal reconstruction were performed, 10 of which were jejunoplasties: 5 free jejunums and 5 supercharged. Morbidity, mortality, mean stay and withdrawal time from enteral feeding were lower in free than in supercharged jejunums. CONCLUSIONS: Supercharged jejunoplasty was the last option for reconstruction of the thoracic esophagus. Median sternotomy access provides an excellent approach to the anterior mediastinum and the internal mammary vessels. The free jejunum would be the first choice, with the indemnity of the rest of the esophagus, in the reconstruction of the cervical esophagus.

Anterior Interosseous Nerve to Ulnar Nerve Transfer: A Systematic Review.

Background: Ulnar nerve injuries, especially high (proximal forearm) injuries, result in poor functional recovery. Peripheral nerve transfers have recently become a popular technique to augment nerve repairs and reduce the reinnervation distance before distal motor endplates irreversibly degenerate, leading to incomplete recovery. Objectives: To systematically review and analyse the recent literature regarding anterior interosseous nerve (AIN) to ulnar nerve transfers, including demographics, indications, outcomes, and complications. Methods: A search was performed using PubMed, MEDLINE, EMBASE, CINAHL, Scopus, and Cochrane databases using the keywords ulnar nerve, ulnar nerve injury, ulnar motor nerve, anterior interosseous nerve, anterior interosseous, AIN, nerve transfer, and end-to-side using a 3-component search along with the Boolean operators 'AND' and 'OR'. Results: A total of 341 studies were retrieved using the search criteria. Sixteen studies met the inclusion criteria including 12 retrospective case series, 3 retrospective cohort studies, and a single randomised control trial. Nine studies involved supercharged end-to-side transfer (SETS), 6 involved end-to-end transfer (ETE), and only 1 study compared results between SETS and ETE transfers. A total of 269 patients underwent nerve transfers. In the ETE subgroup, the average time to nerve transfer was 7 months, with a mean follow-up period of 24.5 months. Post-procedure, 100% (37/37) patients recovered intrinsic function of BMRC ≥1, and the average recovery time was 3.6 months. A total of 85% of patients recovered intrinsic function of BMRC ≥3. In the SETS group, the average time to nerve transfer was 2.5 months. The average follow-up in this cohort was 13.2 months. About 93% (145/156) recovered the intrinsic function of BMRC ≥1, and the average time to recovery was 7 months. About 75% of patients recovered the intrinsic function of BMRC ≥3 in their first dorsal interossei. Conclusion: AIN to ulnar nerve transfer carries low morbidity, and there is low quality evidence to suggest recovery of intrinsic muscle function compared with conventional primary repair techniques. The supercharged end-to-side transfer (SETS) seems to be more favourable compared with end-to-side transfer. Outcome measurements are highly variable amongst studies, making standardisation difficult. Results of further trials are highly anticipated in this exciting field of peripheral nerve surgery.

The Gantzer transfer - Assessment of the feasibility of using the nerve supplying the Gantzer muscle for end-to-side supercharging of the ulnar nerve.

Our study aimed at assessing the anatomical feasibility of using the nerve supplying the Gantzer muscle (GM) to supercharge the ulnar nerve following injury. The GM nerve was dissected and measured in 36 forearms. The distance between its origin and the lateral epicondyle of humerus and between the GM nerve and the ulnar nerve was measured. The GM was present in 15 forearms (47%). The average distance between the origin of the GM nerve and the lateral epicondyle was 7.34 cm (range 3.3-9.1 cm). The average length of the GM nerve was 3.05 cm (range 1.6-4.5 cm) from origin to neuromuscular junction. The average distance from the ulnar nerve was 2.56 cm (range 1.8-13 3.4 cm). The length of the GM nerve was significantly greater (p < 0.05) than the perpendicular distance between its origin and the ulnar nerve, allowing ample margin for side-to-side or end-to-side supercharging of the ulnar nerve with minimal or no need for further translocation or dissection. The use of the GM nerve as donor following ulnar nerve injury may provide an alternative to the pronator quadratus nerve for supercharged end-to-side transfer, or as an addition, thus supercharging the ulnar nerve twice.

Role of Ionic Strength in the Formation of Stable Supramolecular Nanoparticle-Protein Conjugates for Biosensing.

Monolayer-protected gold nanoparticles (AuNPs) exhibit distinct physical and chemical properties depending on the nature of the ligand chemistry. A commonly employed NP monolayer comprises hydrophobic molecules linked to a shell of PEG and terminated with functional end group, which can be charged or neutral. Different layers of the ligand shell can also interact in different manners with proteins, expanding the range of possible applications of these inorganic nanoparticles. AuNP-fluorescent Green Fluorescent Protein (GFP) conjugates are gaining increasing attention in sensing applications. Experimentally, their stability is observed to be maintained at low ionic strength conditions, but not at physiologically relevant conditions of higher ionic strength, limiting their applications in the field of biosensors. While a significant amount of fundamental work has been done to quantify electrostatic interactions of colloidal nanoparticle at the nanoscale, a theoretical description of the ion distribution around AuNPs still remains relatively unexplored. We perform extensive atomistic simulations of two oppositely charged monolayer-protected AuNPs interacting with fluorescent supercharged GFPs co-engineered to have complementary charges. These simulations were run at different ionic strengths to disclose the role of the ionic environment on AuNP-GFP binding. The results highlight the capability of both AuNPs to intercalate ions and water molecules within the gold-sulfur inner shell and the different tendency of ligands to bend inward allowing the protein to bind not only with the terminal ligands but also the hydrophobic alkyl chains. Different binding stability is observed in the two investigated cases as a function of the ligand chemistry.

Improved transfer efficiency of supercharged 36 + GFP protein mediate nucleic acid delivery.

The potential of nucleic acid therapeutics to treat diseases by targeting specific cells has resulted in its increasing number of uses in clinical settings. However, the major challenge is to deliver bio-macromolecules into target cells and/or subcellular locations of interest ahead in the development of delivery systems. Although, supercharged residues replaced protein 36 + GFP can facilitate itself and cargoes delivery, its efficiency is still limited. Therefore, we combined our recent progress to further improve 36 + GFP based delivery efficiency. We found that the penetration efficacy of 36 + GFP protein was significantly improved by fusion with CPP-Dot1l or treatment with penetration enhancer dimethyl sulfoxide (DMSO) in vitro. After safely packaged with plasmid DNA, we found that the efficacy of in vitro and in vivo transfection mediated by 36 + GFP-Dot1l fusion protein is also significantly improved than 36 + GFP itself. Our findings illustrated that fusion with CPP-Dot1l or incubation with DMSO is an alternative way to synergically promote 36 + GFP mediated plasmid DNA delivery in vitro and in vivo.

Differences in Tumor Growth and Differentiation in NSG and Humanized-BLT Mice; Analysis of Human vs. Humanized-BLT-Derived NK Expansion and Functions.

There is significant interest and debate regarding the best mouse model of human disease, since studies in wild-type mice may not always recapitulate human diseases. The NSG mouse model has been one of the most commonly used mouse models to study cancer; however, this mouse model, even though it has several advantages in regard to the ease of tumor implantation and financial feasibility, does not represent human disease due to the immunodeficient nature of this model. In this study, we performed oral and pancreatic tumor studies in NSG and hu-BLT mice and found several distinguishing features that make hu-BLT model more suitable for studying human cancer. In addition, we compared the immune function of humans to hu-BLT mice to understand the differences and similarities of the models. Oral and pancreatic cancer stem cells were implanted in NSG and hu-BLT mice. Both tumors grew robustly in NSG mice and killed them within a short period of time. On the contrary, unlike NSG mice, tumor-bearing hu-BLT mice survived longer, grew smaller tumors, and the grown tumors exhibited lower rates of expansion, with a higher surface expression of MHC-class I and lower NK cell-mediated cytotoxicity that was previously shown to have more of a differentiated phenotype. Although the peripheral blood of hu-BLT mice in comparison to that of humans had lower percentages of NK cells and cytotoxic function, it mediated a higher secretion of IFN-γ, likely contributing to the differentiation of the tumor cells and subsequent decrease in the tumor size in the hu-BLT mice in comparison to the NSG mice. Spleen-derived hu-BLT mouse NK cells were able to expand in the presence of autologous osteoclasts and substantially increase both cytotoxicity and secretion of IFN-γ, similar to those seen in peripheral blood-derived human NK cells, indicating that NK cells from hu-BLT mice are capable of expansion and functional activation when activating signals are given. Thus, the many similarities between human and hu-BLT mouse immune systems make this mouse model more appropriate to study human cancer. In particular, it is well-suited for studies of allogeneic NK cell-based immunotherapy in cancer treatment. The advantages and challenges of hu-BLT mice in cancer studies are also discussed in this report.

Ulnar nerve decompression and transposition with versus without supercharged end-to-side motor nerve transfer for advanced cubital tunnel syndrome: a randomized comparison study.

OBJECTIVE: The objective of this prospective randomized study was to compare ulnar nerve decompression and anterior subfascial transposition with versus without supercharged end-to-side anterior interosseous nerve-to-ulnar motor nerve transfer for advanced cubital tunnel syndrome, to describe performing the nerve transfer through a small incision, and to investigate predictive factors for poor recovery following the procedure. METHODS: Between January 2013 and October 2016, 93 patients were randomly allocated to a study group (n = 45) and a control group (n = 48). Patients in the study group were treated with supercharged motor nerve transfer via a 5-cm incision following decompression and anterior subfascial transposition. Patients in the control group were treated with decompression and anterior subfascial transposition alone. Postoperative pinch strength and compound muscle action potential amplitude (CMAPa) were assessed. Function of the limb was assessed based on the Gabel/Amadio scale. Between-group data were compared, and significance was set at p < 0.05. Potential risk factors were collected from demographic data and disease severity indicators. RESULTS: At the final follow-up at 2 years, the results of the study group were superior to those of the control group with regard to postoperative pinch strength (75.13% ± 7.65% vs 62.11% ± 6.97%, p < 0.05); CMAPa of the first dorsal interossei (17.17 ± 5.84 mV vs 12.20 ± 4.09 mV, p < 0.01); CMAPa of abductor digiti minimi (11.57 ± 4.04 mV vs 8.43 ± 6.11 mV, p < 0.01); and excellent to good results (0.67 for the study group vs 0.35 for the control group, p < 0.05). Multivariate analysis showed that the advanced age (OR 2.98, 95% CI 2.25-4.10; p = 0.003) in the study group was related to unsatisfactory outcome in the patients. CONCLUSIONS: In the treatment of advanced cubital tunnel syndrome, additional supercharged end-to-side anterior interosseous nerve-to-ulnar motor nerve transfer may produce a better function of the hand. The authors also found that cases in the elderly were related to unsatisfactory postoperative results for these patients and that they could be informed of the possibility of worsening surgery results.

Utilizing the ABC Transporter for Growth Factor Production by fleQ Deletion Mutant of Pseudomonas fluorescens.

Pseudomonas fluorescens, a gram-negative bacterium, has been proven to be a capable protein manufacturing factory (PMF). Utilizing its ATP-binding cassette (ABC) transporter, a type I secretion system, P. fluorescens has successfully produced recombinant proteins. However, besides the target proteins, P. fluorescens also secretes unnecessary background proteins that complicate protein purification and other downstream processes. One of the background proteins produced in large amounts is FliC, a flagellin protein. In this study, the master regulator of flagella gene expression, fleQ, was deleted from P. fluorescens Δtp, a lipase and protease double-deletion mutant, via targeted gene knockout. FleQ directs flagella synthesis, so the new strain, P. fluorescens ΔfleQ, does not produce flagella-related proteins. This not only simplifies purification but also makes P. fluorescens ΔfleQ an eco-friendly expression host because it will not survive outside a controlled environment. Six recombinant growth factors, namely, insulin-like growth factors I and II, beta-nerve growth factor, fibroblast growth factor 1, transforming growth factor beta, and tumor necrosis factor beta, prepared using our supercharging method, were successfully secreted by P. fluorescens ΔfleQ. Our findings demonstrate the potential of P. fluorescens ΔfleQ, combined with our supercharging process, as a PMF.