Thermographic Changes During Soft Tissue Incisions with Diode Infrared Lasers.

Objective: This study aimed to demonstrate temperature changes and heat transfer patterns in soft tissues when using infrared (IR) diode lasers, utilizing thermographic techniques. Methods: Bovine tongue slices (5 mm thick) were placed between two glass slides at 11 cm from a thermographic camera. Twenty-two centimeter-long incisions were made along the soft tissue parallel to the camera capture field. Incisions were performed using the 970 and 980 nm lasers (continuous wave, 2-watt, 320 µm-thick glass initiated, and noninitiated fiber tips, 30-sec irradiation). The maximum temperature changes in oC (ΔT) and the vertical and lateral heat transfer (in mm) were recorded for 30 sec, using the thermographic images captured using the IR camera. The ΔT and the amount of lateral and vertical heat distribution were measured in 10-sec intervals for a 30-sec irradiation period. A repeated analysis of variance (ANOVA) (p < 0.05) statistical test was used to analyze the statistical differences between the average ΔT and heat transfer patterns between the initiated and noninitiated lasers. Results: The maximum ΔT for the 970 nm diode laser with initiated tips at the 30-sec mark was 17.81 ± 11.48, while the maximum ΔT for the 980 nm diode laser with initiated tips was 13.24 ± 6.90 (p = 0.041). Statistically significant differences between the vertical and horizontal heat transfer patterns were noted between the initiated and noninitiated diode lasers. The 980 nm diode laser with initiated tips proved to have statistically significant greater vertical and lateral heat transfer when compared to the 970 nm diode laser. The 970 nm diode laser with noninitiated tips proved to have a statistically significant higher heat distribution when compared to the 980 nm laser with noninitiated tips. Conclusions: Different near-IR lasers present differences in lateral heat and tissue penetration, using initiated or noninitiated fibers, and due to these differences, power settings and irradiation period must be considered to avoid risks due to overheating.

Reconceptualizing Family Engagement as an Improvisational Practice: Lessons from Pre-K Teachers' Practices During COVID-19.

Scholars have argued against a post-COVID return to normality on the grounds that the pandemic offers an opportunity to break with the past and imagine a different, more just future. In this analysis of pre-kindergarten teachers' reflections on teaching during COVID-19 in the state of Michigan, we take up the notion of the pandemic as a portal to consider how practices that emerged during the pandemic might be carried forward post-pandemic. Through a qualitative interview study with 25 public pre-K teachers in Michigan, we sought to understand how the pandemic altered the nature of family-teacher engagement. Our analysis led us to conceptualize teaching as an improvisational practice that was highly responsive to the circumstances and needs of families. We identified three central themes that animated pre-K teachers' work during the pandemic: supporting families through new types of "offers" (a term from improv theory), making learning accessible, and fostering collectivity by partnering with families. Teachers' practices during the pandemic reveal new avenues for conceptualizing family engagement as an improvisational practice. We draw on the principles of improv theory to outline a framework for this approach.

Nanoparticles Bind to Endothelial Cells in Injured Blood Vessels via a Transient Protein Corona.

Nanoparticles travel through blood vessels to reach disease sites, but the local environment they encounter may affect their surface chemistry and cellular interactions. Here, we found that as nanoparticles transit through injured blood vessels they may interact with a highly localized concentration of platelet factor 4 proteins released from activated platelets. The platelet factor 4 binds to the nanoparticle surface and interacts with heparan sulfate proteoglycans on endothelial cells, and induces uptake. Understanding nanoparticle interactions with blood proteins and endothelial cells during circulation is critical to optimizing their design for diseased tissue targeting and delivery.

Resolving Dimensionality in a Child Assessment Tool: An Application of the Multilevel Bifactor Model.

Multidimensionality and hierarchical data structure are common in assessment data. These design features, if not accounted for, can threaten the validity of the results and inferences generated from factor analysis, a method frequently employed to assess test dimensionality. In this article, we describe and demonstrate the application of the multilevel bifactor model to address these features in examining test dimensionality. The tool for this exposition is the Child Observation Record Advantage 1.5 (COR-Adv1.5), a child assessment instrument widely used in Head Start programs. Previous studies on this assessment tool reported highly correlated factors and did not account for the nesting of children in classrooms. Results from this study show how the flexibility of the multilevel bifactor model, together with useful model-based statistics, can be harnessed to judge the dimensionality of a test instrument and inform the interpretability of the associated factor scores.

A proposed mathematical description of in vivo nanoparticle delivery.

Nanoparticles are promising vehicles for the precise delivery of molecular therapies to diseased sites. Nanoparticles interact with a series of tissues and cells before they reach their target, which causes less than 1% of administered nanoparticles to be delivered to these target sites. Researchers have been studying the nano-bio interactions that mediate nanoparticle delivery to develop guidelines for designing nanoparticles with enhanced delivery properties. In this review article, we describe these nano-bio interactions with a series of mathematical equations that quantitatively define the nanoparticle delivery process. We employ a compartment model framework to describe delivery where nanoparticles are either (1) at the site of administration, (2) in the vicinity of target cells, (3) internalized by the target cells, or (4) sequestered away in off-target sites or eliminated from the body. This framework explains how different biological processes govern nanoparticle transport between these compartments, and the role of intercompartmental transport rates in determining the final nanoparticle delivery efficiency. Our framework provides guiding principles to engineer nanoparticles for improved targeted delivery.

Identifying cell receptors for the nanoparticle protein corona using genome screens.

Nanotechnology provides platforms to deliver medical agents to specific cells. However, the nanoparticle's surface becomes covered with serum proteins in the blood after administration despite engineering efforts to protect it with targeting or blocking molecules. Here, we developed a strategy to identify the main interactions between nanoparticle-adsorbed proteins and a cell by integrating mass spectrometry with pooled genome screens and Search Tool for the Retrieval of Interacting Genes analysis. We found that the low-density lipoprotein (LDL) receptor was responsible for approximately 75% of serum-coated gold nanoparticle uptake in U-87 MG cells. Apolipoprotein B and complement C8 proteins on the nanoparticle mediated uptake through the LDL receptor. In vivo, nanoparticle accumulation correlated with LDL receptor expression in the organs of mice. A detailed understanding of how adsorbed serum proteins bind to cell receptors will lay the groundwork for controlling the delivery of nanoparticles at the molecular level to diseased tissues for therapeutic and diagnostic applications.

A 3D culture platform enables development of zinc-binding prodrugs for targeted proliferation of ß cells.

Advances in treating ß cell loss include islet replacement therapies or increasing cell proliferation rate in type 1 and type 2 diabetes, respectively. We propose developing multiple proliferation-inducing prodrugs that target high concentration of zinc ions in ß cells. Unfortunately, typical two-dimensional (2D) cell cultures do not mimic in vivo conditions, displaying a markedly lowered zinc content, while 3D culture systems are laborious and expensive. Therefore, we developed the Disque Platform (DP)-a high-fidelity culture system where stem cell-derived ß cells are reaggregated into thin, 3D discs within 2D 96-well plates. We validated the DP against standard 2D and 3D cultures and interrogated our zinc-activated prodrugs, which release their cargo upon zinc chelation-so preferentially in ß cells. Through developing a reliable screening platform that bridges the advantages of 2D and 3D culture systems, we identified an effective hit that exhibits 2.4-fold increase in ß cell proliferation compared to harmine.

An Analysis of the Binding Function and Structural Organization of the Protein Corona.

Blood proteins adsorb onto the surface of nanoparticles after intravenous injection to form a protein corona. The underlying organization and binding function of these adsorbed proteins remain unclear. This can impact how the corona mediates cell and tissue interactions. Here, we investigated the function and structural organization of the protein corona using an immunoassay approach. We discovered that only 27% of the adsorbed proteins examined are functional for binding to their target protein. This is because the corona architecture is not a monolayer, but an assembly of proteins that are bound to each other. We further demonstrated that we can control the binding functionality of a protein by changing the organization of proteins in the assembly. We show that manipulation of the corona protein composition and assembly can influence their interactions with macrophage cells in culture. This study provides detailed functional and structural insights into the protein corona on nanomaterials and offers a new strategy to manipulate it for controlled interactions with the biological system.

Liposome Imaging in Optically Cleared Tissues.

Three-dimensional (3D) optical microscopy can be used to understand and improve the delivery of nanomedicine. However, this approach cannot be performed for analyzing liposomes in tissues because the processing step to make tissues transparent for imaging typically removes the lipids. Here, we developed a tag, termed REMNANT, that enables 3D imaging of organic materials in biological tissues. We demonstrated the utility of this tag for the 3D mapping of liposomes in intact tissues. We also showed that the tag is able to monitor the release of entrapped therapeutic agents. We found that liposomes release their cargo >100-fold faster in tissues in vivo than in conventional in vitro assays. This allowed us to design a liposomal formulation with enhanced ability to kill tumor associated macrophages. Our development opens up new opportunities for studying the chemical properties and pharmacodynamics of administered organic materials in an intact biological environment. This approach provides insight into the in vivo behavior of degradable materials, where the newly discovered information can guide the engineering of the next generation of imaging and therapeutic agents.

The entry of nanoparticles into solid tumours.

The concept of nanoparticle transport through gaps between endothelial cells (inter-endothelial gaps) in the tumour blood vessel is a central paradigm in cancer nanomedicine. The size of these gaps was found to be up to 2,000 nm. This justified the development of nanoparticles to treat solid tumours as their size is small enough to extravasate and access the tumour microenvironment. Here we show that these inter-endothelial gaps are not responsible for the transport of nanoparticles into solid tumours. Instead, we found that up to 97% of nanoparticles enter tumours using an active process through endothelial cells. This result is derived from analysis of four different mouse models, three different types of human tumours, mathematical simulation and modelling, and two different types of imaging techniques. These results challenge our current rationale for developing cancer nanomedicine and suggest that understanding these active pathways will unlock strategies to enhance tumour accumulation.

Rapid mixing of colliding picoliter liquid droplets delivered through-space from piezoelectric-actuated pipettes characterized by time-resolved fluorescence monitoring.

Rapid mixing of aqueous solutions is a crucial first step to study the kinetics of fast biochemical reactions with high temporal resolution. Remarkable progress toward this goal has been made through the development of advanced stopped-flow mixing techniques resulting in reduced dead times, and thereby extending reaction monitoring capabilities to numerous biochemical systems. Concurrently, piezoelectric actuators for through-space liquid droplet sample delivery have also been applied in several experimental systems, providing discrete picoliter sample volume delivery and precision sample deposition onto a surface, free of confinement within microfluidic devices, tubing, or other physical constraints. Here, we characterize the inertial mixing kinetics of two aqueous droplets (130 pl) produced by piezoelectric-actuated pipettes, following droplet collision in free space and deposition on a surface in a proof of principle experiment. A time-resolved fluorescence system was developed to monitor the mixing and fluorescence quenching of 5-carboxytetramethylrhodamine (5-Tamra) and N-Bromosuccinimide, which we show to occur in less than 10 ms. In this respect, this methodology is unique in that it offers millisecond mixing capabilities for very small quantities of discrete sample volumes. Furthermore, the use of discrete droplets for sample delivery and mixing in free space provides potential advantages, including the elimination of the requirement for a physical construction as with microfluidic systems, and thereby makes possible and extends the experimental capabilities of many systems.

Characterizing the protein corona of sub-10 nm nanoparticles.

Studies into the interactions of serum proteins with nanoparticles are typically performed using nanoparticles that are larger than the size of proteins. Due to this size discrepancy, adsorbed proteins are commonly depicted as a globular structure surrounding a nanoparticle. Here, we asked how we should view nanoparticle-protein complexes when the nanoparticles are of similar size or smaller than the proteins with which they interact. We showed that nanoparticles can serve as a cargo on a protein rather than as a carrier of the protein in a size-dependent manner. This can occur when nanoparticles are below 10 nm in diameter. We discovered that when the nanoparticle is a cargo on the protein, the binding of the protein to the receptor target is minimally affected in contrast to the nanoparticle serving as a carrier. Our study should change how we view and describe nanoparticle-protein complexes when the nanoparticles involved are equal in size or smaller than proteins.

Elimination Pathways of Nanoparticles.

Understanding how nanoparticles are eliminated from the body is required for their successful clinical translation. Many promising nanoparticle formulations for in vivo medical applications are large (>5.5 nm) and nonbiodegradable, so they cannot be eliminated renally. A proposed pathway for these nanoparticles is hepatobiliary elimination, but their transport has not been well-studied. Here, we explored the barriers that determined the elimination of nanoparticles through the hepatobiliary route. The route of hepatobiliary elimination is usually through the following pathway: (1) liver sinusoid, (2) space of Disse, (3) hepatocytes, (4) bile ducts, (5) intestines, and (6) out of the body. We discovered that the interaction of nanoparticles with liver nonparenchymal cells ( e. g., Kupffer cells and liver sinusoidal endothelial cells) determines the elimination fate. Each step in the route contains cells that can sequester and chemically or physically alter the nanoparticles, which influences their fecal elimination. We showed that the removal of Kupffer cells increased fecal elimination by >10 times. Combining our results with those of prior studies, we can start to build a systematic view of nanoparticle elimination pathways as it relates to particle size and other design parameters. This is critical to engineering medically useful and translatable nanotechnologies.

beta2 Integrins are characteristically absent in acute promyelocytic leukemia and rapidly upregulated in vivo upon differentiation with all-trans retinoic acid.

Although little is known about migration of hematopoietic stem cells and their neoplastic counterparts into tissues and peripheral blood, adhesion proteins likely play an important role. We studied 339 patients with acute myelogenous leukemia (AML) to discern the relationship between adhesion protein expression, circulating blasts, and white blood cell (WBC) count. Expression levels of CD11b and CD11c strongly correlated with increased WBC count, independent of FAB subtype (p<0.0001). However, 93% (25/27) of cases of AML-M3 completely lacked beta2 integrin expression, compared to 11% (35/312) of the non-M3 cases (p<0.0001). Seven of the 27 patients with AML-M3 were followed during standard induction therapy with ATRA. Within 3 days, weak CD11c became detectable, followed by CD11b and CD11a. Our data suggest an important link between beta2 integrin expression and the level of circulating leukemic cells in AML. We demonstrate the clinical usefulness of a panel of beta2 integrins (CD11a, CD11b and CD11c) in accurate prediction of AML-M3, and recommend inclusion of this immunophenotypic analysis to identify patients who require ATRA therapy. Finally, we illustrate the rapidity at which AML-M3 blasts up-regulate beta2 integrins, and suggest a possible association between this finding and the tissue infiltration that characterizes the "ATRA syndrome".

Hamartin and tuberin immunohistochemical expression in cutaneous fibroepithelial polyps.

BACKGROUND: Hamartin and tuberin are inactivating tumor suppressor proteins implicated in the development of gastrointestinal polyps and sporadic and tuberous sclerosis-associated cutaneous angiofibromas. The pattern of expression of these peptides has not been studied in fibroepithelial polyps (FEPs). DESIGN: The specific aim of the study was to evaluate the immunohistochemical expression of tuberin and hamartin within the epithelium and dermal fibrocytes of 20 cutaneous FEPs compared with the epithelium and dermal fibrocytes of normal skin. The diagnoses were confirmed independently by a dermatopathologist, and the pattern of intensity was assessed by the mean labeling intensity (MLI) of cytoplasmic and/or nuclear staining for each antibody. RESULTS: Hamartin and tuberin antibodies showed moderate staining of the keratinocytes and fibrocytes of normal skin and the keratinocytes within FEPs. Both antibodies showed diminished staining within the fibrocytes of the FEPs. The MLI of hamartin was 44.3 +/- 4.4 for keratinocyte nuclei in normal skin and 51.2 +/- 3.7 within the polyps. The MLI of tuberin was 42.9 +/- 3.6 within the keratinocyte nuclei of the normal skin compared to 39.7 +/- 3.0 for the polyps. The MLI for hamartin within the fibrocytes of the normal skin was 78.9 +/- 7.1 compared to 21.6 +/- 4.2 within the polyps, p = 0.01. The MLI for tuberin within the fibrocytes of normal skin was 70.6 +/- 5.0 compared to 47.1 +/- 4.7 within the polyps. CONCLUSION: The data suggest that down regulation or loss of tuberin and/or hamartin expression may be permissive to fibrocyte proliferation or promote collagen production leading to FEP formation.