Off-axis holographic laser speckle contrast imaging of blood vessels in tissues

Laser speckle contrast imaging (LSCI) has become one of the most common tools for functional imaging in tissues. Incomplete theoretical description and sophisticated interpretation of measurement results are completely sidelined by a low-cost and simple hardware, fastness, consistent results, and repeatability. In addition to the relatively low measuring volume with around 700   μ m of the probing depth for the visible spectral range of illumination, there is no depth selectivity in conventional LSCI configuration; furthermore, in a case of high NA objective, the actual penetration depth of light in tissues is greater than depth of field (DOF) of an imaging system. Thus, the information about these out-of-focus regions persists in the recorded frames but cannot be retrieved due to intensity-based registration method. We propose a simple modification of LSCI system based on the off-axis holography to introduce after-registration refocusing ability to overcome both depth-selectivity and DOF problems as well as to get the potential possibility of producing a cross-section view of the specimen

WPI Hydrogels with a Prolonged Drug-Release Profile for Antimicrobial Therapy

Infectious sequelae caused by surgery are a significant problem in modern medicine due to their reduction of therapeutic effectiveness and the patients’ quality of life.Recently, new methods of local antimicrobial prophylaxis of postoperative sequelae have been actively developed. They allow high local concentrations of drugs to be achieved, increasing the antibiotic therapy’s effectiveness while reducing its side effects. We have developed and characterized antimicrobial hydrogels based on an inexpensive and biocompatible natural substance from the dairy industry—whey protein isolate—as matrices for drug delivery. The release of cefazolin from the pores of hydrogel structures directly depends on the amount of the loaded drug and occurs in a prolonged manner for three days. Simultaneously with the antibiotic release, hydrogel swelling and partial degradation occurs. The WPI hydrogels absorb solvent, doubling in size in three days and retaining cefazolin throughout the duration of the experiment. The antimicrobial activity of cefazolin-loaded WPI hydrogels against Staphylococcus aureus growth is prolonged in comparison to that of the free cefazolin. The overall cytotoxic effect of cefazolin-containing WPI hydrogels is lower than that of free antibiotics. Thus, our work shows that antimicrobial WPI hydrogels are suitable candidates for local antibiotic therapy of infectious surgical sequelae

pH-sensitive dairy-derived hydrogels with a prolonged drug release profile for cancer treatment

A novel versatile biocompatible hydrogel of whey protein isolate (WPI) and two types of tannic acid (TAs) was prepared by crosslinking of WPI with TAs in a one-step method at high temperature for 30 min. WPI is one common protein-based preparation which is used for hydrogel formation. The obtained WPI-TA hydrogels were in disc form and retained their integrity after sterilization by autoclaving. Two TA preparations of differing molecular weight and chemical structure were compared, namely a polygalloyl glucose-rich extract-ALSOK 02-and a polygalloyl quinic acid-rich extract-ALSOK 04. Hydrogel formation was observed for WPI solutions containing both preparations. The swelling characteristics of hydrogels were investigated at room temperature at different pH values, namely 5, 7, and 9. The swelling ability of hydrogels was independent of the chemical structure of the added TAs. A trend of decrease of mass increase (MI) in hydrogels was observed with an increase in the TA/WPI ratio compared to the control WPI hydrogel without TA. This dependence (a MI decrease-TA/WPI ratio) was observed for hydrogels with different types of TA both in neutral and acidic conditions (pH 5.7). Under alkaline conditions (pH 9), negative values of swelling were observed for all hydrogels with a high content of TAs and were accompanied by a significant release of TAs from the hydrogel network. Our studies have shown that the release of TA from hydrogels containing ALSOK04 is higher than from hydrogels containing ALSOK 02. Moreover, the addition of TAs, which display a strong anti-cancer effect, increases the cytotoxicity of WPI-TAs hydrogels against the Hep-2 human laryngeal squamous carcinoma (Hep-2 cells) cell line. Thus, WPI-TA hydrogels with prolonged drug release properties and cytotoxicity effect can be used as anti-cancer scaffolds

Optical coherence microangiography of the mouse kidney for diagnosis of circulatory disorders

Optical coherence tomography (OCT) has become widespread in clinical applications in which precise three-dimensional functional imaging of living organs is required. Nevertheless, the kidney is inaccessible for the high resolution OCT imaging due to a high light attenuation coefficient of skin and soft tissues that significantly limits the penetration depth of the probing laser beam. Here, we introduce a surgical protocol and fixation scheme that enables functional visualization of kidney’s peritubular capillaries via OCT microangiography. The model of reversible/irreversible glomerulus embolization using drug microcarriers confirms the ability of OCT to detect circulatory disorders. This approach can be used for choosing optimal carriers, their dosages and diagnosis of other blood flow pathologies

Lightsheet-based flow cytometer for whole blood with the ability for the magnetic retrieval of objects from the blood flow

Detection and extraction of circulating tumor cells and other rare objects in the bloodstream are of great interest for modern diagnostics, but devices that can solve this problem for the whole blood volume of laboratory animals are still rare. Here we have developed SPIM-based lightsheet flow cytometer for the detection of fluorescently-labeled objects in whole blood. The bypass channel between two blood vessels connected with the external flow cell was used to visualize, detect, and magnetically separate fluorescently-labeled objects without hydrodynamic focusing. Carriers for targeted drug delivery were used as model objects to test the device performance. They were injected into the bloodstream of the rat, detected fluorescently, and then captured from the bloodstream by a magnetic separator prior to filtration in organs. Carriers extracted from the whole blood were studied by a number of in vitro methods

Off-axis holographic laser speckle contrast imaging of blood vessels in tissues

Laser speckle contrast imaging (LSCI) has become one of the most common tools for functional imaging in tissues. Incomplete theoretical description and sophisticated interpretation of measurement results are completely sidelined by a low-cost and simple hardware, fastness, consistent results, and repeatability. In addition to the relatively low measuring volume with around 700   μ m of the probing depth for the visible spectral range of illumination, there is no depth selectivity in conventional LSCI configuration; furthermore, in a case of high NA objective, the actual penetration depth of light in tissues is greater than depth of field (DOF) of an imaging system. Thus, the information about these out-of-focus regions persists in the recorded frames but cannot be retrieved due to intensity-based registration method. We propose a simple modification of LSCI system based on the off-axis holography to introduce after-registration refocusing ability to overcome both depth-selectivity and DOF problems as well as to get the potential possibility of producing a cross-section view of the specimen

Additive Manufacturing of Drug-Eluting Multilayer Biodegradable Films

Drug-eluting films made of bioresorbable polymers are a widely used tool of modern personalized medicine. However, most currently existing methods of producing coatings do not go beyond the laboratory, as they have low encapsulation efficiency and/or difficulties in scaling up. The PLACE (Printed Layered Adjustable Cargo Encapsulation) technology proposed in this article uses an additive approach for film manufacturing. PLACE technology is accessible, scalable, and reproducible in any laboratory. As a demonstration of the technology capabilities, we fabricated layered drug-eluting polyglycolic acid films containing different concentrations of Cefazolin antibiotic. The influence of the amount of loaded drug component on the film production process and the release kinetics was studied. The specific loading of drugs was significantly increased to 200–400 µg/cm2 while maintaining the uniform release of Cefazolin antibiotic in a dosage sufficient for local antimicrobial therapy for 14 days. The fact that the further increase in the drug amount results in the crystallization of a substance, which can lead to specific defects in the cover film formation and accelerated one-week cargo release, was also shown, and options for further technology development were proposed

Hidden stage of intracranial hemorrhage in newborn rats studied with laser speckle contrast imaging and wavelets

Using the laser speckle contrast imaging and wavelet-based analyses, we investigate a latent (a "hidden") stage of the development of intracranial hemorrhages (ICHs) in newborn rats. We apply two measures based on the continuous wavelet-transform of blood flow velocity in the sagittal sinus, namely, the spectral energy in distinct frequency ranges and a multiscality degree characterizing complexity of experimental data. We show that the wavelet-based multifractal formalism reveals changes in the cerebrovascular blood flow at the development of ICH

Ellipticity imaging for visualizing and quantifying long and short range correlations in laser speckle data II: phantom and animal studies

We present an alternative approach for using dynamic laser speckle data to quantify biophysical dynamics including ordered flows and random motions. The approach yields images that superficially resemble traditional laser speckle contrast images, but instead of relying on the statistics of the local time integrated intensity values calculated over temporal and sliding spatial windows as is done in LSCI to create images, ellipticity imaging (EI) directly yields images that quantify the relative dominance of long-range correlations in the temporal dimension of a series of speckle patterns to the short-range correlations in the same dimension. The approach relies on a Poincaré analysis of the speckle data which yields metrics that statistically describe both the short-terms variations in the temporal speckle intensity (i.e., the standard deviation in successive differences) and also the corresponding long term variations. These metrics are plotted against each other (Poincaré plots) and an ellipse fit to the data. The ratio of the semi-major axis to the semi-minor axis of this ellipse for each temporal speckle sequence is then used as the data to form the images (thus the term EI). The results of flow phantom and mouse EI studies will be presented. Various flow rates of dilute intralipid were illuminated with a coherent laser source and EI images were generated. The same speckle data were analyzed using spatial and temporal LSCI approaches. Flows in anesthetized mouse brain vessels were analyzed using EI and LSCI approaches. The results of the studies using the different speckle analyses will be discussed