Owing to its tumor tropism and prolonged transgene expression, mesenchymal stem cell (MSC) has been considered as an ideal delivery vehicle for cancer gene therapies or therapeutic vaccines. In this ...study, we demonstrated that intratumoral (i.t.) injection of MSCs expressing modified interleukin-12 (MSCs/IL-12M) exhibited stronger tumor-specific T-cell responses and antitumor effects as well as more sustained expressions of IL-12 and interferon (IFN)-γ in both sera and tumor sites than did IL-12M-expressing adenovirus (rAd/IL-12M) in mice bearing both solid and metastatic tumors. Subcutaneous (s.c.) injection of MSCs/IL-12M at contralateral site of tumor exhibited similar levels of serum IL-12 and IFN-γ as i.t. injection, but much weaker antitumor effects in both B16F10 melanoma and TC-1 cervical cancer models than i.t. injection. Although intravenous (i.v.) injection elicited earlier peak serum levels of cytokines, it induced weaker tumor-specific T-cell responses and antitumor effects than i.t. injection, indicating that serum cytokine levels are not surrogate indicators of antitumor effects. Taken together, these results indicated that MSC is more efficient than adenovirus as a cytokine gene delivery vehicle and that i.t. injection of MSCs/IL-12M is the best approach to induce strong tumor-specific T-cell responses that correlate with anti-metastatic effects as well as inhibition of solid tumor growth, although MSCs themselves have an ability to migrate into the tumor site. In addition, MSCs/IL-12M embedded in Matrigel (MSCs/IL-12M/Matrigel) exhibited significant antitumor effects even in immunodeficient mice such as SCID and BNX mice lacking T, B and natural killer (NK) cells, but not in IFN-γ knockout mice. Our findings provide an optimal approach for designing an efficient clinical protocol of MSC-based cytokine gene therapy to induce strong tumor-specific T-cell responses and therapeutic anticancer efficacy.
Propionibacterium acnes is a key therapeutic target in acne, yet this bacterium has become resistant to standard antibiotic agents. We investigated whether the human antimicrobial protein granulysin ...is a potential candidate for the treatment of acne. Granulysin and synthetic granulysin-derived peptides possessing a helix–loop–helix motif killed P. acnes in vitro. Modification of a helix–loop–helix peptide, 31–50, by substitution of a tryptophan for the valine at amino acid 44 (peptide 31–50v44w) to increase its interaction with bacterial surfaces also increased its antimicrobial activity. Moreover, when synthesized with D- rather than L-type amino acids, this peptide (D-31–50v44w) became less susceptible to degradation by proteases and more effective in killing P. acnes. Granulysin peptides were bactericidal, demonstrating an advantage over standard bacteriostatic antibiotics in their control of P. acnes. Moreover, peptide D-31–50v44w killed P. acnes in isolated human microcomedone preparations. Importantly, peptides 31–50, 31–50v44w, and D-31–50v44w also have potential anti-inflammatory effects, as demonstrated by suppression of P. acnes-stimulated cytokine release. Taken together, these data suggest that granulysin peptides may be useful as topical therapeutic agents, providing alternatives to current acne therapies.
Abstract
Background
Our previous study using mesenchymal stem cells (MSCs) as delivery vehicles for tumor necrosis factor-related apoptosis inducing ligand (TRAIL) showed an anti-tumor effect in ...vitro and in vivo for glioblastoma. However, because most malignant glioma cells exhibit TRAIL resistance, we need to find new strategies to overcome it.
We developed MSCs expressing both TRAIL and cytosine deaminase (CD) which convert the prodrug 5-fluorocytosine (5-FC) to the cytotoxic agent 5-fluorouracil (5-FU). We also immortalized these MSCs and introduced a doxycycline-dependent gene-inducible system to maintain the quality control. Lastly, we enhanced tumor tropism by overexpression of CCR2 and CXCR4. In this study, we investigated the anti-tumor efficacy of our novel MSCs (BM-03) with a 5-FC in vitro and in vivo orthotopic xenograft glioblastoma model.
Material and Methods
Human bone marrow-derived MSCs were isolated and cultured as described previously after approval by our institutional review board. The multiple genes mentioned above were transfected into the MSCs using a lentivirus vector system. Using a U87 human glioma cell line and male athymic nude mice, we developed an orthotopic glioblastoma xenograft model. We injected BM-03 in a dose-dependent manner (control; low dose, 2.0X104 cells; intermediate dose, 1.0X105 cells, and high dose, 5.0X105 cells) intratumorally with the intraperitoneal injection of 5-FC (500 mg/kg). Additionally, with in vivo bioluminescence imaging, we assessed the tumor volume.
Results
First, we confirmed the doxycycline-dependent inducible gene expression system of BM-03 and showed the high expression of TRAIL, CCR2, and CXCR4 using FACS analysis. The synergistic anti-tumor efficacy of BM-03 with 5-FC was shown compared with BM-03 alone, in vitro. Moreover, we revealed the enhanced tumor tropism of BM-03 using in vivo bioluminescence imaging. After BM-03 injection of the contralateral side of the tumor, we could see that our MSCs moved toward the inoculated tumor lesion located on the contralateral side. Finally, when treated with BM-03 and 5-FC, we showed an increased antitumor efficacy in terms of both the median survival rate (control, 44 days; low dose, 46 days; intermediate, 57 days, and high dose, 69 days) and the tumor volume in a dose-dependent manner compared with the control and BM-03 only in the orthotopic xenograft glioblastoma mouse model.
Conclusion
Our novel immortalized and inducible MSCs expressing TRAIL and CD showed an enhanced antitumor efficacy in an orthotopic xenograft glioblastoma model. This successful preclinical result will serve as a foundation for IND approval and a phase I clinical trial using our novel MSCs (BM-03).
We describe the framework for capturing the internal view of complex anatomical spaces via multiple media and haptic platforms, exemplified by realistic and conceptual representations of the ...pterygopalatine fossa (PPF). A realistic three-dimensional (3D) mesh of the PPF was developed by segmenting the osseous anatomy on computed tomography (CT) using
Materialize InPrint
. Subsequently in
Autodesk 3D Studio Max
, the realistic mesh was enhanced with graphically designed neurovascular anatomy and additionally a conceptual representation of the PPF with its connections and contents was created. An interactive web-compatible
Adobe Flash
tutorial using
ActionScript
was developed, allowing users to advance through a series of educational slides that contained interactive rotatable interior camera views and scrollable CT cross-sectional content, incorporating both the realistic and conceptual models. Both models were also 3D printed using polyamide material. In the realistic model, the neurovasculature was colored with water-based acrylic paint. A 3-piece modular design with embedded magnets allows for internal visualization and seamless assembly. A serious gaming environment of the conceptual PPF was also developed using
Truevision3D
application programming interface, where users can freely move around rooms and hallways that represent various spaces. Lastly, the realistic model was incorporated into a headset-based virtual reality environment,
Surgical Theater
, allowing visualization and fly-through inside and outside the model. Multiple 3D techniques for visualization of complex 3D anatomical spaces from within were described, with the necessary software and skills detailed. A rough estimate of the time and cost needed to develop these tools as well as multiple supplementary source and end result files are also made available. Educators could utilize multiple advanced delivery methods to incorporate custom digital 3D models of complex anatomical spaces understood from inside.
Mixed‐metal oxyhydroxides—especially those of Ni and Fe—are one of the most active classes of materials known for catalyzing the oxygen evolution reaction (OER). Here, nanoparticulate mixed metal ...oxyhydroxides (of Ni, Fe, and Co) were prepared on an electrode surface by electrochemical reaction of a precursor solution encapsulated in aqueous nanodroplets (AnDs), with each of the droplets containing 10 s of attoliters of fluid. Electrode reactions and synthesis can be monitored in situ by electrochemistry as single AnD stochastically lands and interacts with the working electrode. Resultant metal oxyhydroxide nanoparticles can be size and composition controlled precisely by modulating the precursor solution stored in the AnD. Nanoparticulate metal oxyhydroxides were implemented as catalysts for the OER and exhibited superior catalysis compared to their thin‐film counterparts, demonstrating a hundred‐thousand‐fold enhancement in atom efficiency at comparable turnover rates.
Electrode materials: Stoichiometry‐controlled nanoparticulate mixed‐metal (Co, Fe, Ni) oxyhydroxides were prepared electrochemically by electrolysis of aqueous nanodroplets (AnDs), each containing 10 s of attoliters of fluid. The prepared particles exhibited homogeneous distribution of component metals without phase segregation, and demonstrated superior catalytic activity for the oxygen evolution reaction compared to the thin film counterparts of the same composition (see figure).
Clinical trials of gene therapy using a viral delivery system for glioma have been limited. Recently, gene therapy using stem cells as the vehicles for delivery of therapeutic agents has emerged as a ...new treatment strategy for malignant brain tumors. In this study, we used human umbilical cord blood-derived mesenchymal stem cells (UCB-MSCs) as delivery vehicles with glioma-targeting capabilities, and modified interleukin-12 (IL-12p40N220Q; IL-12M) as a novel therapeutic gene. We also engineered UCB-MSCs to secret IL-12M (UCB-MSC-IL12M) via tetrameric cell-permeable peptide (4HP4)-mediated adenoviral transduction. We confirmed the migratory capacity of UCB-MSC-IL12M toward GL26 mouse glioma cells by an in vitro migration assay and in vivo injection of UCB-MSC-IL12M into the ipsilateral hemisphere of implanted gliomas in C57BL/6 mice. In vivo efficacy experiments showed that intratumoral injection of UCB-MSC-IL12M significantly inhibited tumor growth and prolonged the survival of glioma-bearing mice compared with control mice. Antitumor effects were associated with increased local IL-12M levels, followed by interferon-γ secretion and T-cell infiltration in intracranial gliomas, as well as antiangiogenesis. Interestingly, tumor-free mice after UCB-MSC-IL12M treatment were resistant to ipsilateral and contralateral tumor rechallenge, which was closely associated with tumor-specific long-term T-cell immunity. Thus, our results provide the rationale for designing novel experimental protocols to induce long-term antitumor immunity against intracranial gliomas using UCB-MSCs as an effective delivery vehicle for therapeutic cytokines including IL-12M.
Abstract
Background
The 2017 WHO classification of pituitary tumors has revealed “high-risk pituitary neuroendocrine tumors (PitNETs)”, which were known to have high probability for recurrence. Tumor ...invasion was not included in the pathological grading and classification due to frequent lack of proper pathologic assessment. However, it mentioned tumor invasion as an important prognostic feature in identifying clinically aggressive adenomas.
Material and Methods
We performed a retrospective review of a prospectively collected dataset from January 2018 to March 2021. Patient’s clinical presentation, radiologic features, pathologic findings, and clinical outcome were gathered. Inter-group analysis was performed for high-risk versus low-risk tumors, and invasive versus non-invasive tumors.
Results
Among total 116 cases of PitNETs, high-risk and low-risk tumors were identified in 32 and 84 cases, respectively. The inter-group comparison showed no differences in clinical presentation, radiologic features, pathologic findings, and clinical outcomes.Invasive and non-invasive tumors were identified in 49 and 67 cases, respectively. The invasive group tumors were more symptomatic ( 29 (59.2%) vs. 30 (44.8%), p= 0.031), with larger tumor size over 40mm (9 (18.4%) vs. 1 (1.5%), p = 0.002), and more likely to have Knosp grade higher than 3. The gross-total resection was less achievable (7 (14.3%) vs 26 (38.8%), p = 0.007) However, Ki-67 index showed no significantly difference between the invasive group and non-invasive group (2.0 vs 2.0 , p= 0.556).
Conclusion
According to our study results, the pathologic diagnosis of a high-risk tumor does not necessarily seem to properly reflect the clinical aggressiveness. Tumor invasion, however, seems to better represent the aggressive tumors that requires proper and active treatment.
Oblique lateral interbody fusion (OLIF) offers the solution to problems of anterior lumbar interbody fusion (ALIF) and lateral lumbar interbody fusion (LLIF). However, OLIF technique for degenerative ...spinal diseases of elderly patients has been rarely reported. The objective of this study was to determine the clinical and radiological results of OLIF technique for degenerative spinal diseases in patients under or over 65 years of age.
Sixty-three patients who underwent OLIF procedure were enrolled, including 29 patients who were less than 65 years of age and 34 patients who were over 65 years of age. Fusion rate, change of disc height and lumbar lordotic angle, Numeric Rating Scale (NRS), return to daily activity, patient's satisfaction rate (PSR), and Oswestry disability index (ODI) were used to assess clinical and functional outcomes.
The mean NRS scores for back and leg pain decreased, respectively, from 4.6 and 5.9 to 2.3 and 1.8 in the group A (less than 65 years) and from 4.5 and 6.8 to 2.6 and 2.2 in the group B (over 65 years) at the final follow-up period. The mean ODI scores improved from 48.4 to 24.0% in the group A and from 46.5 to 25.2% in the group B at the final follow-up period. In both groups, the NRS and ODI scores significantly changed preoperatively to postoperatively (p < 0.001). However, statistical analysis yielded no significant difference in postoperative NRS/ODI scores between two groups. In both groups, the changes in the disc height, segmental lordosis, and fusion rate between the preoperative and postoperative periods were significant. The amount of change between preoperative and postoperative disc height, segmental lordosis, and whole lumbar lordosis demonstrated significant intergroup differences (p < 0.05). Overall perioperative complications occurred in 8 of 29 (27.6%) patients in the group A and in 10 of 34 (29.4%) patients in the group B. In both groups, the major complication incidence was 0 and 3%, respectively.
Although there was the slightly high incidence of complication associated with high rate of co-morbidities in elderly patients, OLIF for degenerative lumbar diseases in elderly patients showed favorable clinical and radiological outcomes.
Cell-permeable peptides (CPPs) promote the transduction of nonpermissive cells by recombinant adenovirus (rAd) to improve the therapeutic efficacy of rAd. In this study, branched oligomerization of ...CPPs significantly enhanced the transduction of human mesenchymal stem cells (MSCs) by rAd in a CPP type-independent manner. In particular, tetrameric CPPs increased transduction efficiency at 3000-5000-fold lower concentrations than did monomeric CPPs. Although branched oligomerization of CPPs also increases cytotoxicity, optimal concentrations of tetrameric CPPs required for maximum transduction are at least 300-1000-fold lower than those causing 50% cytotoxicity. Furthermore, although only approximately 60% of MSCs were maximally transduced at 500 muM of monomeric CPPs, >95% of MSCs were transduced with 0.1 muM of tetrameric CPPs. Tetrameric CPPs also significantly increased the formation and net surface charge of CPP/rAd complexes, as well as the binding of rAd to cell membranes at a greater degree than did monomeric CPPs, followed by rapid internalization into MSCs. In a critical-size calvarial defect model, the inclusion of tetrameric CPPs in ex vivo transduction of rAd expressing bone morphogenetic protein 2 into MSCs promoted highly mineralized bone formation. In addition, MSCs that were transduced with rAd expressing brain-derived neurotrophic factor in the presence of tetrameric CPPs improved functional recovery in a spinal cord injury model. These results demonstrated the potential for tetrameric CPPs to provide an innovative tool for MSC-based gene therapy and for in vitro gene delivery to MSCs.
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