Dino J. Ravnic

Inflammation-Induced Intussusceptive Angiogenesis in Murine Colitis

Intussusceptive angiogenesis is a morphogenetic process that forms new blood vessels by the division of a single blood vessel into two lumens. Here, we show that this process of intraluminal division participates in the inflammation‐induced neovascularization associated with chemically induced murine colitis. In studies of both acute (4–7 days) and chronic (28–31 days) colitis, intravital microscopy of intravascular tracers demonstrated a twofold reduction in blood flow velocity. In the acute colitis model, the decreased velocity was associated with marked dilatation of the mucosal plexus. In contrast, chronic inflammation was associated with normal caliber vessels and duplication (and triplication) of the quasi‐polygonal mucosal plexus. Scanning electron microscopy (SEM) of intravascular corrosion casts suggested that pillar formation and septation, previously linked to the morphogenetic process of intussusceptive angiogenesis, were present within days of the onset of inflammation. Four weeks after the onset of inflammation, SEM of vascular corrosion casts demonstrated replication of the mucosal plexus without significant evidence of sprouting angiogenesis. These data suggest that mucosal capillaries have comparable aggregate cross‐sectional area in acute and chronic colitis; however, there is a significant increase in functional capillary density in chronic colitis. We conclude that intussusceptive angiogenesis is a fundamental mechanism of microvascular adaptation to prolonged inflammation. Anat Rec, 2010.

Melittin-induced membrane permeability: a nonosmotic mechanism of cell death.

SummaryDerived from honeybees, melittin is a 26-amino acid, α-helical, membrane-attack protein that efficiently kills mammalian cells. To investigate the contribution of colloid-osmotic effects to the mechanism of cell death, we studied the effect of melittin on lymphocyte membrane permeability and cell volumes. Melittin concentrations of 0.5 to 2.0 μM induced release of membrane permeability markers without total disruption of the cell membrane. At these melittin concentrations, electrical-impedance cytometry demonstrated melittin-induced changes in red blood cell volumes (P<0.01), but no change in lymphocyte cell volumes (P>0.05). Streaming video microscopy, obtaining images of melittintreated lymphocytes at 80-ms intervals, demonstrated a loss of optical density (P<0.001) suggesting a flattening of the cell but no significant increase in cell perimeter (P>0.05). Real-time multiparameter flow cytometry of melittin-treated lymphocytes confirmed simultaneous loss of the cytoplasmic marker, calcein, and uptake of the DNA dye, ethidium homodimer, but demonstrated no increase in forward light scatter. Transmission-electron microscopy of melittin-treated lymphocytes showed normal cell volumes but discontinuities in the cell membrane suggesting direct membrane toxicity. We conclude that melittin causes lymphocyte death by a “leaky patch” mechanism that is independent of colloid-osmotic effects.

Structural adaptations in the murine colon microcirculation associated with hapten-induced inflammation

Background: Blood flowing across the vascular endothelium creates wall shear stress, dependent on velocity of flow and vessel geometry, that tends to disrupt lymphocyte–endothelial cell adhesion. Objective: The microcirculation in a murine model of acute colitis was investigated to identify structural adaptations during acute colitis that may facilitate transmigration. Methods: In 2,4,6-trinitrobenzenesulphonic acid-induced acute colitis, the infiltrating cells and colonic microcirculation was investigated by cellular topographic mapping, corrosion casting and three-dimensional scanning electron microscopy (SEM). Colonic blood velocimetry was performed using intravital microscopy. Results: Clinical and histological parameters suggested a peak inflammatory response at 96 h (p<0.001). The infiltrating cells were spatially related to the mucosal capillary plexus by three-dimensional topographic mapping (p<0.001). In normal mice, corrosion casting and three-dimensional SEM showed a polygonal mucosal plexus supplied by ascending arteries and descending veins. After 2,4,6-trinitrobenzenesulphonic acid stimulation, three-dimensional SEM showed preserved branch angles (p = 0.52) and nominal vessel lengths (p = 0.93), but a significantly dilated mucosal capillary plexus (p<0.001). Intravital microscopy of the mucosal plexus showed a greater than twofold decrease in the velocity of flow (p<0.001). Conclusions: The demonstrable slowing of the velocity of flow despite an increase in volumetric flow suggests that these microvascular adaptations create conditions suitable for leucocyte adhesion and transmigration.

Bridging Mucosal Vessels Associated With Rhythmically Oscillating Blood Flow in Murine Colitis

Oscillatory blood flow in the microcirculation is generally considered to be the result of cardiopulmonary influences or active vasomotion. In this report, we describe rhythmically oscillating blood flow in the bridging vessels of the mouse colon that appeared to be independent of known biological control mechanisms. Corrosion casting and scanning electron microscopy of the mouse colon demonstrated highly branched bridging vessels that connected the submucosal vessels with the mucosal plexus. Because of similar morphometric characteristics (19 ± 11 μm vs. 28 ± 16 μm), bridging arterioles and venules were distinguished by tracking fluorescent nanoparticles through the microcirculation using intravital fluorescence videomicroscopy. In control mice, the blood flow through the bridging vessels was typically continuous and unidirectional. In contrast, two models of chemically induced inflammation (trinitrobenzenesulfonic acid and dextran sodium sulfate) were associated with a twofold reduction in flow velocity and the prominence of rhythmically oscillating blood flow. The blood oscillation was characterized by tracking the bidirectional displacement of fluorescent nanoparticles. Space–time plots and particle tracking of the oscillating segments demonstrated an oscillation frequency between 0.2 and 5.1 cycles per second. Discrete Fourier transforms demonstrated a power spectrum composed of several base frequencies. These observations suggest that inflammation‐inducible changes in blood flow patterns in the murine colon resulted in both reduced blood flow velocity and rhythmic oscillations within the bridging vessels of the mouse colon. Anat Rec, 291:74–82, 2007.

Multiframe particle tracking in intravital imaging: defining Lagrangian coordinates in the microcirculation

The cellular composition of the microcirculation creates blood flow that can be unsteady and nonuniform. To obtain information about nonuniform cellular trajectories, we describe in vivo imaging techniques that provide both detailed tracking of individual particles as well as an approach to simultaneous multicolor particle tracking. Particularly relevant to biologic systems, Lagrangian methods provide information about the fate of individual particles and flow in the system.

Breast implant-associated anaplastic large cell lymphoma: A systematic review

Importance Breast implant–associated anaplastic large cell lymphoma (BIA-ALCL), a rare peripheral T-cell lymphoma, is increasing in incidence. However, many practitioners who treat patients with breast cancer are not aware of this disease. Objectives To assess how BIA-ALCL develops, its risk factors, diagnosis, and subsequent treatment and to disseminate information about this entity to the medical field. Evidence Review A literature review was performed in an academic medical setting. All review articles, case reports, original research articles, and any other articles relevant to BIA-ALCL were included. Data on BIA-ALCL, such as pathophysiology, patient demographics, presentation, diagnosis, treatment, and outcomes, were extracted. Particular focus was paid to age, time to onset, implant type, initial symptoms, treatment, and survival. The search was conducted in January 2017 for studies published in any year. Findings After duplicates were excluded, 304 relevant articles were assessed, and 115 were included from the first documented case in August 1997 through January 2017. Thirty review articles, 44 case reports or series, 15 original research articles, and 26 “other” articles (eg, techniques, special topics, letters) were reviewed. A total of 93 cases have been reported in the literature, and with the addition of 2 unreported cases from the Penn State Health Milton S. Hershey Medical Center, 95 patients were included in this systematic review. Almost all documented BIA-ALCL cases have been associated with a textured device. The underlying mechanism is thought to be due to chronic inflammation from indolent infections, leading to malignant transformation of T cells that are anaplastic lymphoma kinase (ALK) negative and CD30 positive. The mean time to presentation is approximately 10 years after implant placement, with 55 of 83 (66%) patients initially seen with an isolated late-onset seroma and 7 of 83 (8%) with an isolated new breast mass. Ultrasonography with fluid aspiration can be used for diagnosis. Treatment must include removal of the implant and surrounding capsule. More advanced disease may require chemotherapy, radiotherapy, and lymph node dissection. Conclusions and Relevance Breast implant–associated anaplastic large cell lymphoma is a rare cancer in patients with breast implants but is increasing in incidence. It is important for all physicians involved in the care of patients with breast implants to be aware of this entity and be able to recognize initial symptoms.

Transplantation of Bioprinted Tissues and Organs: Technical and Clinical Challenges and Future Perspectives

&NA; Three-dimensional (3D) bioprinting is a revolutionary technology in building living tissues and organs with precise anatomic control and cellular composition. Despite the great progress in bioprinting research, there has yet to be any clinical translation due to current limitations in building human-scale constructs, which are vascularized and readily implantable. In this article, we review the current limitations and challenges in 3D bioprinting, including in situ techniques, which are one of several clinical translational models to facilitate the application of this technology from bench to bedside. A detailed discussion is made on the technical barriers in the fabrication of scalable constructs that are vascularized, autologous, functional, implantable, cost-effective, and ethically feasible. Clinical considerations for implantable bioprinted tissues are further expounded toward the correction of end-stage organ dysfunction and composite tissue deficits.

Bimodal Oscillation Frequencies of Blood Flow in the Inflammatory Colon Microcirculation

Rhythmic changes in blood flow direction have been described in the mucosal plexus of mice with acute colitis. In this report, we studied mice with acute colitis induced either by dextran sodium sulfate or by trinitrobenzenesulfonic acid. Both forms of colitis were associated with blood flow oscillations as documented by fluorescence intravital videomicroscopy. The complex oscillation patterns suggested more than one mechanism for these changes in blood flow. By tracking fluorescent nanoparticles in the inflamed mucosal plexus, we identified two forms of blood flow oscillations within the inflammatory mouse colon. Stable oscillations were associated with a base frequency of approximately 2 cycles/sec. Velocity measurements in the upstream and downstream vessel segments indicated that stable oscillations were the result of regional flow occlusion within the mucosal plexus. In contrast, metastable oscillations demonstrated a lower frequency (0.2–0.4 cycles/sec) and appeared to be the result of flow dynamics in vessels linked by the bridging mucosal vessels. These blood flow oscillations were not directly associated with cardiopulmonary movement. We conclude that both the stable and metasable oscillating patterns reflect flow adaptations to inflammatory changes in the mucosal plexus. Anat Rec, 2009.

Concise Review: Bioprinting of Stem Cells for Transplantable Tissue Fabrication

Bioprinting is a quickly progressing technology, which holds the potential to generate replacement tissues and organs. Stem cells offer several advantages over differentiated cells for use as starting materials, including the potential for autologous tissue and differentiation into multiple cell lines. The three most commonly used stem cells are embryonic, induced pluripotent, and adult stem cells. Cells are combined with various natural and synthetic materials to form bioinks, which are used to fabricate scaffold‐based or scaffold‐free constructs. Computer aided design technology is combined with various bioprinting modalities including droplet‐, extrusion‐, or laser‐based bioprinting to create tissue constructs. Each bioink and modality has its own advantages and disadvantages. Various materials and techniques are combined to maximize the benefits. Researchers have been successful in bioprinting cartilage, bone, cardiac, nervous, liver, and vascular tissues. However, a major limitation to clinical translation is building large‐scale vascularized constructs. Many challenges must be overcome before this technology is used routinely in a clinical setting. Stem Cells Translational Medicine 2017;6:1940–1948

Exploration of small RNA-seq data for small non-coding RNAs in Human Colorectal Cancer

Background: Improved healthcare and recent breakthroughs in technology have substantially reduced cancer mortality rates worldwide. Recent advancements in next-generation sequencing (NGS) have allowed genomic analysis of the human transcriptome. Now, using NGS we can further look into small non-coding regions of RNAs (sncRNAs) such as microRNAs (miRNAs), Piwi-interacting-RNAs (piRNAs), long non-coding RNAs (lncRNAs), and small nuclear/nucleolar RNAs (sn/snoRNAs) among others. Recent studies looking at sncRNAs indicate their role in important biological processes such as cancer progression and predict their role as biomarkers for disease diagnosis, prognosis, and therapy. Results: In the present study, we data mined publically available small RNA sequencing data from colorectal tissue samples of eight matched patients (benign, tumor, and metastasis) and remapped the data for various small RNA annotations. We identified aberrant expression of 13 miRNAs in tumor and metastasis specimens [tumor vs benign group (19 miRNAs) and metastasis vs benign group (38 miRNAs)] of which five were upregulated, and eight were downregulated, during disease progression. Pathway analysis of aberrantly expressed miRNAs showed that the majority of miRNAs involved in colon cancer were also involved in other cancers. Analysis of piRNAs revealed six to be over-expressed in the tumor vs benign cohort and 24 in the metastasis vs benign group. Only two piRNAs were shared between the two cohorts. Examining other types of small RNAs [sn/snoRNAs, mt_rRNA, miscRNA, nonsense mediated decay (NMD), and rRNAs] identified 15 sncRNAs in the tumor vs benign group and 104 in the metastasis vs benign group, with only four others being commonly expressed. Conclusion: In summary, our comprehensive analysis on publicly available small RNA-seq data identified multiple differentially expressed sncRNAs during colorectal cancer progression at different stages compared to normal colon tissue. We speculate that deciphering and validating the roles of sncRNAs may prove useful in colorectal cancer prognosis, diagnosis, and therapy.