Histochemical Staining of Collagen and Identification of Its Subtypes by  Picrosirius Red Dye in Mouse Reproductive Tissues

Bhutda, Smita; Surve, Manalee Vishnu; Anil, Anjali; Kamath, Kshama Ganesh; Singh, Neha; Modi, Deepak; Banerjee, Anirban

doi:10.21769/bioprotoc.2592

Cited by 23 publications

(20 citation statements)

References 9 publications

(1 reference statement)

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“…The local area was calculated near the center of mass of MNP distribution with a radius equals to the maximum value of the interquartile ranges of the MNP distributions in the vertical and horizontal directions. Cytoplasm area was quantified as the yellow area in PR stained sections 64 , which was extracted from the “b” channel in the “l, a, b” color space. Tissue area was computed via binary thresholding, which excluded the dark-brown areas that represented MNPs and the white background.…”

Section: Methodsmentioning

confidence: 99%

Biomechanical sensing of in vivo magnetic nanoparticle hyperthermia-treated melanoma using magnetomotive optical coherence elastography

et al. 2021

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Rationale: Magnetic nanoparticle hyperthermia (MH) therapy is capable of thermally damaging tumor cells, yet a biomechanically-sensitive monitoring method for the applied thermal dosage has not been established. Biomechanical changes to tissue are known indicators for tumor diagnosis due to its association with the structural organization and composition of tissues at the cellular and molecular level. Here, by exploiting the theranostic functionality of magnetic nanoparticles (MNPs), we aim to explore the potential of using stiffness-based metrics that reveal the intrinsic biophysical changes of in vivo melanoma tumors after MH therapy. Methods: A total of 14 melanoma-bearing mice were intratumorally injected with dextran-coated MNPs, enabling MH treatment upon the application of an alternating magnetic field (AMF) at 64.7 kHz. The presence of the MNP heating sources was detected by magnetomotive optical coherence tomography (MM-OCT). For the first time, the elasticity alterations of the hyperthermia-treated, MNP-laden, in vivo tumors were also measured with magnetomotive optical coherence elastography (MM-OCE), based on the mechanical resonant frequency detected. To investigate the correlation between stiffness changes and the intrinsic biological changes, histopathology was performed on the excised tumor after the in vivo measurements. Results: Distinct shifts in mechanical resonant frequency were observed only in the MH-treated group, suggesting a heat-induced stiffness change in the melanoma tumor. Moreover, tumor cellularity, protein conformation, and temperature rise all play a role in tumor stiffness changes after MH treatment. With low cellularity, tumor softens after MH even with low temperature elevation. In contrast, with high cellularity, tumor softening occurs only with a low temperature rise, which is potentially due to protein unfolding, whereas tumor stiffening was seen with a higher temperature rise, likely due to protein denaturation. Conclusions: This study exploits the theranostic functionality of MNPs and investigates the MH-induced stiffness change on in vivo melanoma-bearing mice with MM-OCT and MM-OCE for the first time. It was discovered that the elasticity alteration of the melanoma tumor after MH treatment depends on both thermal dosage and the morphological features of the tumor. In summary, changes in tissue-level elasticity can potentially be a physically and physiologically meaningful metric and integrative therapeutic marker for MH treatment, while MM-OCE can be a suitable dosimetry technique.

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Section: Methodsmentioning

confidence: 99%

Biomechanical sensing of in vivo magnetic nanoparticle hyperthermia-treated melanoma using magnetomotive optical coherence elastography

et al. 2021

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“…We further utilized the ARS and MP slides for semi-quantitative analysis of calcium and elastin contents, respectively. As in previous studies, [ 30 , 31 , 32 ], the elastin and calcium quantifications were performed as follows:

…”

Section: Methodsmentioning

confidence: 99%

Animal Model Dependent Response to Pentagalloyl Glucose in Murine Abdominal Aortic Injury

Anderson

Niedert

Patnaik

et al. 2021

JCM

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Abdominal aortic aneurysms (AAAs) are a local dilation of the aorta and are associated with significant mortality due to rupture and treatment complications. There is a need for less invasive treatments to prevent aneurysm growth and rupture. In this study, we used two experimental murine models to evaluate the potential of pentagalloyl glucose (PGG), which is a polyphenolic tannin that binds to and crosslinks elastin and collagen, to preserve aortic compliance. Animals underwent surgical aortic injury and received 0.3% PGG or saline treatment on the adventitial surface of the infrarenal aorta. Seventeen mice underwent topical elastase injury, and 14 mice underwent topical calcium chloride injury. We collected high-frequency ultrasound images before surgery and at 3–4 timepoints after. There was no difference in the in vivo effective maximum diameter due to PGG treatment for either model. However, the CaCl2 model had significantly higher Green–Lagrange circumferential cyclic strain in PGG-treated animals (p < 0.05). While ex vivo pressure-inflation testing showed no difference between groups in either model, histology revealed reduced calcium deposits in the PGG treatment group with the CaCl2 model. These findings highlight the continued need for improved understanding of PGG’s effects on the extracellular matrix and suggest that PGG may reduce arterial calcium accumulation.

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“…When visualised using bright field microscopy collagen appears as bright pink to red bundles and the cells (nuclei) exhibit shades of brown. 35 Briefly, the tissue sections were dewaxed in xylene, hydrated using descending series of ethanol followed by a dip in distilled water. The sections were immersed in hematoxylin for half an hour, followed by a dip in running water and then immersed in ammonia solution for 10 s. The sections were then stained with picrosirius stain (sirius red stain prepared in picric acid solution) for 1 h, followed by a dip in acid water for 2 s and were then subjected to ascending series of ethanol.…”

Section: Methodsologymentioning

confidence: 99%

Altered presence of extra cellular matrix components in murine skin cancer: Modulation by Azadirachta indica leaf extract

Chugh

Koul

2021

Journal of Traditional and Complementary Medicine

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Background and aim Although, the anticancer potential of Aqueous Azadirachta indica leaf extract (AAILE) has been robustly established against cutaneous squamous cell carcinoma (SCC) in mice, however, its ability in modulating tumor associated extra cellular matrix (ECM) is largely unknown. Therefore, the present study was conceived to explore changes in ECM during murine skin cancer and its chemoprevention by AAILE. Experimental procedure Skin tumors were induced using a two-stage model of carcinogenesis employing topical application of 7,12-Dimethylbenz(a)anthracene (DMBA) and 12-O-tetradecanoyl phorbol-13-acetate (TPA) as carcinogen and promoter respectively. AAILE was administered orally to the animals. Male Laca mice were divided into four groups: control, AAILE, DMBA/TPA and AAILE + DMBA/TPA. Results The tumors obtained in DMBA/TPA and AAILE + DMBA/TPA groups were histologically identified as SCC. Tumor induction in these groups was accompanied by raised serum carcinoembryonic antigen (CEA) levels when compared to control counterparts. Assessment of hydroxyproline levels and histochemical staining with sirius red and trichrome stain revealed an increase in collagen in tumors of DMBA/TPA group. An increase in glycosaminoglycans (GAGs) levels was also observed in DMBA/TPA group as made evident by biochemical studies and histochemical staining using mucicarmine and alcian blue-periodic acid schiff’s stain. Administration of AAILE to DMBA/TPA treated animals caused a decrease in collagen and GAG levels along with a decrease in serum CEA levels. Conclusion Skin tumors exhibited altered presence of ECM components which is indicative of a modified ECM. AAILE administration antagonised tumor associated ECM alterations which may be contributing to its chemopreventive activity as reported previously.

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Histochemical Staining of Collagen and Identification of Its Subtypes by Picrosirius Red Dye in Mouse Reproductive Tissues

Cited by 23 publications

References 9 publications

Biomechanical sensing of in vivo magnetic nanoparticle hyperthermia-treated melanoma using magnetomotive optical coherence elastography

Biomechanical sensing of in vivo magnetic nanoparticle hyperthermia-treated melanoma using magnetomotive optical coherence elastography

Animal Model Dependent Response to Pentagalloyl Glucose in Murine Abdominal Aortic Injury

Altered presence of extra cellular matrix components in murine skin cancer: Modulation by Azadirachta indica leaf extract

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