scholarly journals Dantrolene Prevents the Lymphostasis Caused by Doxorubicin in the Rat Mesenteric Circulation

2021 ◽  
Vol 12 ◽  
Author(s):  
Serena Van ◽  
Soumiya Pal ◽  
Brittney R. Garner ◽  
Kate Steed ◽  
Vijayalakshmi Sridharan ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
High Speed ◽  
Ryanodine Receptors ◽  
Lymph Flow ◽  
Free Calcium ◽  
Physiological Salt Solution ◽  
Breast Cancer Patients ◽  
Rapid Infusion ◽  
Prophylactic Use

Background and Purpose: Doxorubicin (DOX) is a risk factor for arm lymphedema in breast cancer patients. We reported that DOX opens ryanodine receptors (RYRs) to enact “calcium leak,” which disrupts the rhythmic contractions of lymph vessels (LVs) to attenuate lymph flow. Here, we evaluated whether dantrolene, a clinically available RYR1 subtype antagonist, prevents the detrimental effects of DOX on lymphatic function.Experimental Approach: Isolated rat mesenteric LVs were cannulated, pressurized (4–5 mm Hg) and equilibrated in physiological salt solution and Fura-2AM. Video microscopy recorded changes in diameter and Fura-2AM fluorescence tracked cytosolic free calcium ([Ca2+i]). High-speed in vivo microscopy assessed mesenteric lymph flow in anesthetized rats. Flow cytometry evaluated RYR1 expression in freshly isolated mesenteric lymphatic muscle cells (LMCs).Key Results: DOX (10 μmol/L) increased resting [Ca2+i] by 17.5 ± 3.7% in isolated LVs (n = 11). The rise in [Ca2+i] was prevented by dantrolene (3 μmol/L; n = 10). A single rapid infusion of DOX (10 mg/kg i.v.) reduced positive volumetric lymph flow to 29.7 ± 10.8% (n = 7) of baseline in mesenteric LVs in vivo. In contrast, flow in LVs superfused with dantrolene (10 μmol/L) only decreased to 76.3 ± 14.0% (n = 7) of baseline in response to DOX infusion. Subsequently, expression of the RYR1 subtype protein as the presumed dantrolene binding site was confirm in isolated mesenteric LMCs by flow cytometry.Conclusion and Implications: We conclude that dantrolene attenuates the acute impairment of lymph flow by DOX and suggest that its prophylactic use in patients subjected to DOX chemotherapy may lower lymphedema risk.

scholarly journals Dynamic blood flow phantom for in vivo liquid biopsy standardization

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Anastasiia Kozlova ◽  
Daniil Bratashov ◽  
Oleg Grishin ◽  
Arkadii Abdurashitov ◽  
Ekaterina Prikhozhdenko ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Blood Cells ◽  
Liquid Biopsy ◽  
High Speed ◽  
White Blood Cells ◽  
High Sensitivity ◽  
Acoustic Properties ◽  
Detection Methods ◽  
In Vivo Flow Cytometry

AbstractIn vivo liquid biopsy, especially using the photoacoustic (PA) method, demonstrated high clinical potential for early diagnosis of deadly diseases such as cancer, infections, and cardiovascular disorders through the detection of rare circulating tumor cells (CTCs), bacteria, and clots in the blood background. However, little progress has been made in terms of standardization of these techniques, which is crucial to validate their high sensitivity, accuracy, and reproducibility. In the present study, we addressed this important demand by introducing a dynamic blood vessel phantom with flowing mimic normal and abnormal cells. The light transparent silica microspheres were used as white blood cells and platelets phantoms, while hollow polymeric capsules, filled with hemoglobin and melanin, reproduced red blood cells and melanoma CTCs, respectively. These phantoms were successfully used for calibration of the PA flow cytometry platform with high-speed signal processing. The results suggest that these dynamic cell flow phantoms with appropriate biochemical, optical, thermal, and acoustic properties can be promising for the establishment of standardization tool for calibration of PA, fluorescent, Raman, and other detection methods of in vivo flow cytometry and liquid biopsy.

scholarly journals In vivo photothermal flow cytometry: Imaging and detection of individual cells in blood and lymph flow

2006 ◽  
Vol 97 (5) ◽  
pp. 916-932 ◽  
Author(s):  
Vladimir P. Zharov ◽  
Ekaterina I. Galanzha ◽  
Valery V. Tuchin
Keyword(s):  
Flow Cytometry ◽  
Lymph Flow

scholarly journals ALG3 contributes to stemness and radioresistance through regulating glycosylation of TGF-β receptor II in breast cancer

2021 ◽  
Vol 40 (1) ◽  
Author(s):  
Xiaoqing Sun ◽  
Zhenyu He ◽  
Ling Guo ◽  
Caiqin Wang ◽  
Chuyong Lin ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Flow Cytometry ◽  
Cancer Patients ◽  
Early Recurrence ◽  
Breast Cancer Patients ◽  
Rt Pcr ◽  
Cancer Stemness ◽  
Β Receptor

Abstract Background Radiotherapy is a conventional and effective local treatment for breast cancer. However, residual or recurrent tumors appears frequently because of radioresistance. Novel predictive marker and the potential therapeutic targets of breast cancer radioresistance needs to be investigated. Methods In this study, we screened all 10 asparagine-linked glycosylation (ALG) members in breast cancer patients’ samples by RT-PCR. Cell viability after irradiation (IR) was determined by CCK-8 assay and flow cytometry. The radiosensitivity of cell lines with different ALG3 expression was determined with the colony formation assay by fitting the multi-target single hit model to the surviving fractions. Cancer stem-like traits were assessed by RT-PCR, Western blot, and flow cytometry. The mechanisms of ALG3 influencing radiosensitivity was detected by Western blot and immunoprecipitation. And the effect of ALG3 on tumor growth after IR was verified in an orthotopic xenograft tumor models. The association of ALG3 with prognosis of breast cancer patients was confirmed by immunohistochemistry. Results ALG3 was the most significantly overexpressing gene among ALG family in radioresistant breast cancer tissue. Overexpression of ALG3 predicted poor clinicopathological characteristics and overall survival (OS), and early local recurrence-free survival (LRFS) in breast cancer patients. Upregulating ALG3 enhanced radioresistance and cancer stemness in vitro and in vivo. Conversely, silencing ALG3 increased the radiosensitivity and repressed cancer stemness in vitro, and more importantly inhibition of ALG3 effectively increased the radiosensitivity of breast cancer cells in vivo. Mechanistically, our results further revealed ALG3 promoted radioresistance and cancer stemness by inducing glycosylation of TGF-β receptor II (TGFBR2). Importantly, both attenuation of glycosylation using tunicamycin and inhibition of TGFBR2 using LY2109761 differentially abrogated the stimulatory effect of ALG3 overexpression on cancer stemness and radioresistance. Finally, our findings showed that radiation played an important role in preventing early recurrence in breast cancer patients with low ALG3 levels, but it had limited efficacy in ALG3-overexpressing breast cancer patients. Conclusion Our results suggest that ALG3 may serve as a potential radiosensitive marker, and an effective target to decrease radioresistance by regulating glycosylation of TGFBR2 in breast cancer. For patients with low ALG3 levels, radiation remains an effective mainstay therapy to prevent early recurrence in breast cancer.

scholarly journals Combined Ultrasound Treatment with Transferrin-Coupled Nanoparticles Improves Active Targeting of 4T1 Mammary Carcinoma Cells

2021 ◽  
Vol 20 ◽  
pp. 153303382110623
Author(s):  
Xiangzi Jin ◽  
Jie Yu ◽  
Meijiao Yin ◽  
Amit Sinha ◽  
Guangming Jin
Keyword(s):  
Breast Cancer ◽  
Flow Cytometry ◽  
Side Effects ◽  
Antitumor Activity ◽  
Breast Cancer Patients ◽  
Growth Inhibitory ◽  
Effective Combination ◽  
Mammary Carcinoma Cells ◽  
Induced Apoptosis

Objective: Conventional chemotherapy remains the mainstay treatment for many breast cancer patients, but its effectiveness is limited by toxic side effects. Incorporating drugs such as docetaxel into nanoparticle medicines can reduce toxicity but further improvements are required. To facilitate more active tumor targeting, we prepared transferrin-docetaxel-loaded pegylated-albumin nanoparticles (Tf-PEG-DANPS). Methods: The growth inhibitory effects and the ability of unmodified DANPS or PEG-DANPS to induce apoptosis in 4T1 mouse mammary cancers were compared to Tf-PEG-DANPS treatment using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and flow cytometry. These experiments were extended in vivo to the intravenous treatment of 4T1 tumors where PEG-DANPS was compared to Tf-PEG-DANPS alone or Tf-PEG-DANPS combined with ultrasound (US + Tf-PEG-DANPS). Histological assessments using hematoxylin and eosin (HE) sections were performed to examine antitumor activity, metastasis to lung and liver, and body weight measurements taken as an indicator of toxicity. Results: MTT experiments show that, in the normal and low concentration interval, the inhibition ability of the Tf-PEG-DANPS is higher than that of other drug-giving groups, and the flow cytometry show that the proportion of induced apoptosis in each given group is 2.88%, 42.95%, 48.23%, and 57.89%, indicating that the Tf-PEG-DANPS group has more significant ability to induce apoptosis than other drug-giving groups. From the pathological HE staining and semiquantitative analysis, US+Tf-PEG-DANPS can effectively inhibit the growth of breast cancer transplanted tumors and suppress metastases, it also has smaller toxic side effects on mice. Conclusion: The antitumor effect of US+Tf-PEG-DANPS represents an effective combination that exhibits increased antitumor activity and metastasis reduction with an improved side-effect profile.

scholarly journals Calcium mobilization in human platelets using indo-1 and flow cytometry

Blood ◽  
1989 ◽  
Vol 74 (8) ◽  
pp. 2674-2680 ◽  
Author(s):  
LK Jennings ◽  
ME Dockter ◽  
CD Wall ◽  
CF Fox ◽  
DM Kennedy
Keyword(s):  
Flow Cytometry ◽  
Flow Cytometric Analysis ◽  
Cytosolic Calcium ◽  
Human Platelets ◽  
Membrane Receptors ◽  
Free Calcium ◽  
Cytoplasmic Calcium ◽  
Calcium Indicator ◽  
Cytoplasmic Free Calcium

Abstract Regulation of cytoplasmic free calcium concentration is believed to be important in the response of platelets to external stimuli. A relatively new fluorescent calcium indicator, indo-1, has properties by which alterations of cytoplasmic calcium can be evaluated in single platelets by flow cytometry. Activation of platelets at a temperature lower than 37 degrees C allows examination of the heterogeneity of intracellular free calcium levels and can distinguish variations among platelets in the initiation, duration, and magnitude of calcium fluxes. The clear advantage of flow cytometric analysis of platelet cytosolic calcium is that stimulus-response coupling can now be studied on a single cell basis. Platelets were activated by addition of human alpha- thrombin or ADP at 37 degrees C or at room temperature (22 degrees C). Activation at 37 degrees C approaches more closely an in vivo response and, as expected, increases in cytosolic calcium occurred within seconds of agonist addition. Transient increases in cytoplasmic calcium levels occurred when platelets were challenged with a low concentration of agonist. Heterogeneity in cytoplasmic calcium levels was also observed at 10(-5) mol/L ADP and 0.1 U/mL alpha-thrombin. Some of this heterogeneity was no longer observed at higher concentrations of agonist (10(-4) mol/L ADP and 0.5 U/mL thrombin), suggesting that a sufficient magnitude of signal is required to induce changes in platelet cytosolic calcium. Light-scatter properties of the activated platelets were also monitored simultaneously and showed changes in response to both agonists. The ability to measure changes in cytoplasmic free calcium by ratio flow cytofluorimetry provides a new approach to study of the role of alterations in intracellular calcium in response to agonists acting through different membrane receptors as well as providing a sensitive technique to detect functional subpopulations of platelets.

scholarly journals Calcium mobilization in human platelets using indo-1 and flow cytometry

Blood ◽  
1989 ◽  
Vol 74 (8) ◽  
pp. 2674-2680
Author(s):  
LK Jennings ◽  
ME Dockter ◽  
CD Wall ◽  
CF Fox ◽  
DM Kennedy
Keyword(s):  
Flow Cytometry ◽  
Flow Cytometric Analysis ◽  
Cytosolic Calcium ◽  
Human Platelets ◽  
Membrane Receptors ◽  
Free Calcium ◽  
Cytoplasmic Calcium ◽  
Calcium Indicator ◽  
Cytoplasmic Free Calcium

Regulation of cytoplasmic free calcium concentration is believed to be important in the response of platelets to external stimuli. A relatively new fluorescent calcium indicator, indo-1, has properties by which alterations of cytoplasmic calcium can be evaluated in single platelets by flow cytometry. Activation of platelets at a temperature lower than 37 degrees C allows examination of the heterogeneity of intracellular free calcium levels and can distinguish variations among platelets in the initiation, duration, and magnitude of calcium fluxes. The clear advantage of flow cytometric analysis of platelet cytosolic calcium is that stimulus-response coupling can now be studied on a single cell basis. Platelets were activated by addition of human alpha- thrombin or ADP at 37 degrees C or at room temperature (22 degrees C). Activation at 37 degrees C approaches more closely an in vivo response and, as expected, increases in cytosolic calcium occurred within seconds of agonist addition. Transient increases in cytoplasmic calcium levels occurred when platelets were challenged with a low concentration of agonist. Heterogeneity in cytoplasmic calcium levels was also observed at 10(-5) mol/L ADP and 0.1 U/mL alpha-thrombin. Some of this heterogeneity was no longer observed at higher concentrations of agonist (10(-4) mol/L ADP and 0.5 U/mL thrombin), suggesting that a sufficient magnitude of signal is required to induce changes in platelet cytosolic calcium. Light-scatter properties of the activated platelets were also monitored simultaneously and showed changes in response to both agonists. The ability to measure changes in cytoplasmic free calcium by ratio flow cytofluorimetry provides a new approach to study of the role of alterations in intracellular calcium in response to agonists acting through different membrane receptors as well as providing a sensitive technique to detect functional subpopulations of platelets.

scholarly journals In vivo multispectral, multiparameter, photoacoustic lymph flow cytometry with natural cell focusing, label-free detection and multicolor nanoparticle probes

2008 ◽  
Vol 73A (10) ◽  
pp. 884-894 ◽  
Author(s):  
Ekaterina I. Galanzha ◽  
Evgeny V. Shashkov ◽  
Valery V. Tuchin ◽  
Vladimir P. Zharov
Keyword(s):  
Flow Cytometry ◽  
Lymph Flow ◽  
Label Free ◽  
Nanoparticle Probes ◽  
Label Free Detection ◽  
Natural Cell

Mitochondrial matrix calcium ions regulate energy production in myocardium: A cytochemical study

1990 ◽  
Vol 48 (2) ◽  
pp. 166-167
Author(s):  
W.A. Jacob ◽  
R. Hertsens ◽  
A. Van Bogaert ◽  
M. De Smet
Keyword(s):  
Energy Metabolism ◽  
Calcium Ions ◽  
Energy Production ◽  
Regulation Of Respiration ◽  
Free Calcium ◽  
Mitochondrial Matrix ◽  
Cytochemical Study ◽  
Nmr Techniques

In the past most studies of the control of energy metabolism focus on the role of the phosphorylation potential ATP/ADP.Pi on the regulation of respiration. Studies using NMR techniques have demonstrated that the concentrations of these compounds for oxidation phosphorylation do not change appreciably throughout the cardiac cycle and during increases in cardiac work. Hence regulation of energy production by calcium ions, present in the mitochondrial matrix, has been the object of a number of recent studies.Three exclusively intramitochondnal dehydrogenases are key enzymes for the regulation of oxidative metabolism. They are activated by calcium ions in the low micromolar range. Since, however, earlier estimates of the intramitochondnal calcium, based on equilibrium thermodynamic considerations, were in the millimolar range, a physiological correlation was not evident. The introduction of calcium-sensitive probes fura-2 and indo-1 made monitoring of free calcium during changing energy metabolism possible. These studies were performed on isolated mitochondria and extrapolation to the in vivo situation is more or less speculative.

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