Cell Proliferation Kinetics Research Articles

DNA Damage, Cell Death, and Alteration of Cell Proliferation Insights Caused by Copper Oxide Nanoparticles Using a Plant-Based Model.

The speedy growth of copper oxide nanoparticle (CuO NP) manufacturing due to their wide application in industries has caused concerns due to their increased discharge into the environment from both purposeful and accidental sources. Their presence at an elevated concentration in the environment can cause potential hazards to the plant kingdom, specifically to staple food crops. However, limited research is available to determine the consequences of CuO NPs. The present study aimed to assess the morphological and cytological changes induced by CuO NPs on Pisum sativum L., a key staple food crop. Seeds of Pisum sativum were exposed to various concentrations of CuO NPs (0, 25, 50, 75, 100, and 125 ppm) for 2 h, and their effects on seed germination (SG), radicle length (RL), cell proliferation kinetics (CPK), mitotic index (MI), cell death (CD), micronucleus frequency (MNF), and chromosomal aberration frequency (CAF) were studied. The results indicate a significant reduction in SG, RL, CPK, and MI and a significant dose-dependent increase in CD, MNF, and CAF. CuO NP treatment has led to abnormal meiotic cell division, increased incidence of micronucleus frequency, and chromosomal aberration frequency. Additionally, the CuO NP-treated groups showed an increase in the percentage of aberrant meiotic cells such as laggard (LG), double bridge (DB), stickiness (STC), clumped nuclei (CNi), precocious separation (PS), single bridge (SB), and secondary association (SA). CuO NP treatment led to reductions in SG as follows: 55% at 24 h, 60.10% at 48 h, and 65% at 72 h; reductions in RL as follows: 0.55 ± 0.021 cm at 24 h, 0.67 ± 0.01 cm at 48 h, and 0.99 ± 0.02 cm at 72 h; reductions in CPK as follows: 34.98% at prophase, 7.90% at metaphase, 3.5% at anaphase, and 0.97% at telophase. It also led to a 57.45% increase in CD, a 39.87% reduction in MI, and a 60.77% increase in MNF at a higher concentration of 125 ppm. The findings of this study clearly show that CuO NPs have a genotoxic effect on the food crop plant Pisum sativum.

Administration sequences in single-day chemotherapy regimens for breast cancer: a comprehensive review from a practical perspective.

Breast cancer is one of the most prevalent malignant tumors globally, posing a severe threat to human life and health. Chemotherapy, a cornerstone in the treatment of breast cancer, often overlooks the sequence of drug administration within single-day regimens. This study aims to explore the impact of drug administration order on the efficacy and toxicity of combination chemotherapy protocols for breast cancer. Through a comprehensive review and analysis based on current evidence from evidence-based medicine, we delved into how the order of drug administration affects both efficacy and toxicity. We systematically classified and analyzed commonly used combination drug regimens, providing graded recommendations and a reasoned analysis to offer valuable references for clinical decision-making. Our findings indicate that the sequence of drug administration in complex combination chemotherapy protocols is not arbitrary but necessitates multifaceted considerations. Rational drug sequencing can maximize synergistic effects between drugs, thereby augmenting therapeutic efficacy while effectively mitigating drug-related adverse effects. Additionally, some drug labels and clinical trials have explicitly highlighted the therapeutic benefits of specific drug sequences. This study underscores the importance of considering the sequence of drug administration in clinical practice. It is recommended to prioritize the sequential drug administration according to official drug product labeling, while also considering factors such as the administration sequence from large randomized controlled trials, cell proliferation kinetics specific to cancer types, drug interactions, chronopharmacology, drug irritability, clinical experiences, and patient preferences. By taking these factors into account, the goal is to maximize treatment efficacy and minimize the occurrence of adverse reactions.

Mathematical Model of Cancer with Ordinary Differential Equations

The cancer cell starts dividing, infiltrating neighboring tissues and traveling throughout the lymphatic system. While there are ways to stop the spread of disease or get rid of infected cells, most of the approaches are unable to identify the early warning indicators of such an occurrence. Using diverse types of differential equations, especially ordinary differential equations (ODEs), is a helpful catalyst that experts employ. Using differential equations, researching resistance to chemotherapy, forecasting potential treatment failure, or evaluating the result and prognosis following various forms of therapy. Living things always include cancer cells, but a biological regulatory system keeps them from spreading to a dangerous degree (think overpopulation vs. natural resources). Therefore, the most efficient method of determining when to effectively intervene with the tumor growth is to use the cytokinetic method of quantitatively assessing the cancer cells’ progression. Cancer Metabolism: One of the main characteristics of cancer is metabolic reprogramming, in which the metabolism of cancer cells is changed to fuel their explosive growth and multiplication. New models of cancer metabolism investigate the roles that dysregulated metabolic pathways play in the genesis and spread of tumors, providing prospective avenues for therapeutic intervention. The mathematical models of tumor growth modeling of ordinary differential equations (ODEs cancer). The tumor grows voraciously, and the scientists and mathematicians who tried to have a better understanding grow. The study of such treatments on models of tumor growth leads to one or more ODEs. Which gives some ideas on the relation between equations and tumor growth in cancer cells introduce ODEs to provide mathematical models of tumor growth. The dynamics of tumor cells and their growths through clinical, experimental, and theoretical approaches, new ideas for different cancer therapies are developed with the goal of controlling and reducing the death rate for earlier diagnosis. The kinetics of tumor cell proliferation and its treatment approach were covered in this research. In order to comprehend the proliferation of tumor cells, we expanded the study and xamined a feew basic mathematical models.

Quantifying Human Naïve B Cell Proliferation Kinetics and Differentiation in Controlled In Vitro Cell Culture.

Division tracking dyes like Cell Trace Violet (CTV) enable the quantification of cell proliferation, division, and survival kinetics of human naïve B cell responses in vitro. Human naïve B cells exhibit distinct responses to different stimuli, with CpG and anti-Ig inducing a T cell-independent (TI) response, while CD40L and IL-21 promote a T cell-dependent (TD) response that induces isotype switching and differentiation into antibody-secreting cells (ASCs). Both stimulation methods yield valuable insights into the intrinsic programming of B cell health within individuals, making them useful for clinical investigations. For instance, quantitative analysis from these B cell populations could reveal biologically meaningful measurements such as the average number of division rounds and the time to cells' fate. Here, we describe a novel in vitro culture setup for CTV-labelled human naïve B cells and a method for obtaining precise time-based data on proliferation, division-linked isotype switching, and differentiation.

Phenthoate toxicity evaluation in root meristem of Pisum sativum L.

Phenthoate is an organothiophosphate insecticide. Effect of phenthoate on the cytogenetic alterations in root tip cells of Pisum sativum L., a multiuse crop was investigated in this study. Pisum sativum L. seeds were exposed to different concentrations of phenthoate (0.1, 0.2, 0.3, 0.4, and 0.5%) and were germinated at 24°C for 72 hours and cytogenetic alterations were assessed. Analysis of mitotic index revealed that phenthoate has cytotoxic attributes, and cell proliferation kinetics frequencies showed alterations in the kinetics of the mitotic process. Phenthoate treatment of 0.1% to 0.5% resulted in an increase in the metaphases, and a reduction in prophases, anaphases, and telophases ratio, dose dependently. The findings of the study reveal that, phenthoate reduced the percentage of seed germination, mitotic index, radicle length and increased chromosomal abnormalities dose dependently. Root tip cells of Pisum sativum L. seeds treated with phenthoate showed an increased occurrence of single and double bridges, fragments, stickiness, laggard, and vagrants.

1723-P: Dual-Label Kinetic Assay to Monitor ß-Cell Death and Proliferation

A deficit of β-cell mass is common to all types of diabetes. Thus, many research efforts are dedicated to developing therapies to preserve and expand β-cell mass. Current methods assessing β-cell proliferation and cell death are limited due to reliance on a single time point, and usually cannot measure proliferation and death simultaneously. Here we have developed and optimized a novel method, based on the single-cell and population-level analyses using real-time kinetic labeling (SPARKL), to study kinetics of cell proliferation and death in β cells. SPARKL uses two fluorescent dyes - SYTO21 labels all cells in green, and YOYO3, a red cell-impermeable viability dye that stains dead cells. We continuously imaged cells every few hours over numerous days and have optimized growth conditions and dye concentrations to monitor INS1 or primary rodent or human islet cells on single cell and population levels. As a first step, we compared growth under 6, 11 and 20 mM glucose and glucolipotoxicity (20 mM glucose and 1 mM palmitate). We found a doubling time of 52±2, 48±4, 44±3 and >120 h, respectively, in INS1 cells. At 48 h, fewer than 16% of the cells are dead in varying glucose concentrations compared to 69±5% dead cells in the glucolipotoxic condition (N=8). Harmine, a known mitogen of β-cells, enhanced proliferation in all glucose concentrations by 5-15% and protected from glucolipotoxic cell death (N=3). In primary islets, as expected, the proliferation rates were extremely low and often masked by cell death in varying glucose concentrations. Glucolipotoxicity led to >80% cell death in mouse and human islets (N=3). Finally, we screened a library of 73 epigenetic modifiers to screen for compounds that rescued INS1 cells from glucolipotoxic-like stress (overexpression of ChREBPβ). Our screen yielded four epigenetic modifiers that reduced cell death to less than 2% at 48 h. In conclusion, SPARKL is a novel and useful method that is accurate, unbiased, and requires minimal handling to simultaneously measure β-cell proliferation and death. Disclosure L.S.Katz: None. P.Wang: None. H.U.Kaniskan: None. J.D.Gelles: None. J.Chipuk: None. A.F.Stewart: None. D.Scott: None. Funding National Institutes of Health (R01DK130300)

Selective Inhibition of L-type Amino Acid Transporter 1 Suppresses Cell Proliferation in Ovarian Clear Cell Carcinoma.

Ovarian clear cell carcinoma (OCCC) is a histological type of ovarian cancer that is refractory to chemotherapy and has poor prognosis, which necessitates the development of novel treatment therapies. In this study, we focused on L-type amino acid transporter 1 (LAT1), which is involved in cancer growth, and investigated the effect of its selective inhibition on cell proliferation in OCCC. The inhibitory effect of nanvuranlat (JPH203), a LAT1 selective inhibitor, on the cellular uptake of [3H] leucine was evaluated using the OCCC cell line JHOC9, which expresses the LAT1 protein. In addition, the kinetics of cell proliferation and changes in phosphorylation of the mTOR pathway were analyzed. The correlation between LAT1 expression and progression-free survival (PFS) was evaluated using clinical specimens of OCCC. Nanvuranlat inhibited [3H] leucine intracellular uptake and cell proliferation in a dose-dependent manner in JHOC9 cells. In addition, it suppressed the activity of the mTOR signaling pathway, which is thought to inhibit cancer cell proliferation. LAT1 expression was most frequent in OCCC among clinical specimens of epithelial ovarian cancer. A correlation between LAT1 expression and PFS was observed in OCCC. LAT1 selective inhibition suppresses cell proliferation via the mTOR pathway by inhibiting leucine uptake in OCCC. This study illustrates the potential of using LAT1 selective inhibition as a treatment strategy for OCCC.

Comprehensive Genomic Profiling Identifies FAT1 as a Negative Regulator of EMT, CTCs, and Metastasis of Hepatocellular Carcinoma.

FAT atypical cadherin 1 (FAT1) acts as a tumor suppressor or oncogene, which regulates cell adherence, proliferation, motility, and actin kinetics. FAT1 gene expression is closely related to hepatocarcinogenesis; however, the function and mechanism of FAT1 in hepatocellular carcinoma (HCC) remain unclear. Here, we screened for the FAT1, which is intimately linked to the development and progression of HCC, both in circulating tumor cells (CTCs) and tumor tissues using next generation sequencing (NGS). Immunohistochemical staining was performed to detect FAT1 protein expression. To determine the impact of FAT1 on epithelial-mesenchymal transition (EMT), migration and invasion of HCC, an in vitrotranswell assay and Western blot were performed. Moreover, Gene Set Enrichment Analysis was carried out to discover the underlying mechanism. Finally, animal experimentswere conducted to confirm the effects of FAT1 on HCC metastasis and tumorigenicity. Our results showed that FAT1 expression was decreased in HCC tissues, while in vitro and in vivo, the FAT1 knockdown facilitated invasion, cell motility, colony formation, and proliferation. FAT1 knockdown also resulted in decreased expression of E-cadherin and markedly elevated expression of N-cadherin, vimentin, and snail. We also confirmed our hypothesis from the analysis of group differences in the CTC phenotype and lung metastasis in nude mice. Our findings illustrated that FAT1 played a negative regulatory role in the HCC EMT and metastasis, providing further evidence for the role played by FAT1 in the formation and progression of HCC.

Methomyl has clastogenic and aneugenic effects and alters the mitotic kinetics in Pisum sativum L.

Methomyl is a carbamate pesticide that is frequently applied to crops all over the world. This research aims to evaluate the Pisum sativum L mitotic process and potential genotoxicity. The Cell Proliferation Kinetics (CPK) frequencies demonstrated changes in kinetics of mitotic process, and study of Mitotic Index (MI) demonstrated that methomyl had cytotoxic properties. In fact, the telophases ratio dropped at 0.1% to 0.5% methomyl treatment, while there was an increase in prophases, metaphases, and anaphases from 0.1% to 0.5% in a dose dependent manner. In terms of genotoxicity, methomyl cause an increase in the frequency of clastogenic and aneugenic chromosomal abnormalities at metaphase-anaphase at 0.1% to 0.5%. The effects on the mitotic spindle were further confirmed by an increase in the frequencies of c-mitosis from 0.1 to 0.5% methomyl treatment. The outcome of the current analysis indicates that regularly used insecticides methomyl has a considerable cytotoxic effect on mitotic cells of Pisum sativum L.

Macroporous 3D printed structures for regenerative medicine applications

The use of natural biopolymers as a core material to produce cell-laden scaffolds has been recognized and extensively utilized for tissue engineering purposes due to their advantageous biocompatibility and tunable biodegradation rate. The morphology and average pore size play, however, a major role in biological processes affecting cell proliferation kinetics as well as tissue regeneration processes associated with extracellular matrix formation. Shear thinning properties of the inks employed in 3D printing for high-accuracy hydrogel scaffold fabrication are often associated with compromises in morphology, such as reduced pore sizes. Here, we report on a carefully optimized composite formulation of (1:1) gelatin/oxidized alginate (Gel/OxAlg) that allows combining 3D printing and cryogelation techniques for simple and low-cost fabrication of biocompatible hydrogel scaffolds, characterized by high porosity and extra-large pore size (d > 100 μm). Based on the morphological characteristics and obtained cell viability data, the fabricated scaffolds might be used as a platform for a variety of tissue engineering applications.

Blood, toil, and taxoteres: Biological determinants of treatment-induced ctDNA dynamics for interpreting tumor response.

Collection and analysis of circulating tumor DNA (ctDNA) is one of the few methods of liquid biopsy that measures generalizable and tumor specific molecules, and is one of the most promising approaches in assessing the effectiveness of cancer care. Clinical assays that utilize ctDNA are commercially available for the identification of actionable mutations prior to treatment and to assess minimal residual disease after treatment. There is currently no clinical ctDNA assay specifically intended to monitor disease response during treatment, partially due to the complex challenge of understanding the biological sources of ctDNA and the underlying principles that govern its release. Although studies have shown pre- and post-treatment ctDNA levels can be prognostic, there is evidence that early, on-treatment changes in ctDNA levels are more accurate in predicting response. Yet, these results also vary widely among cohorts, cancer type, and treatment, likely due to the driving biology of tumor cell proliferation, cell death, and ctDNA clearance kinetics. To realize the full potential of ctDNA monitoring in cancer care, we may need to reorient our thinking toward the fundamental biological underpinnings of ctDNA release and dissemination from merely seeking convenient clinical correlates.

Studies on Cytotoxic and Genotoxic potential of Ethanolic extract of Lawsonia inermis leaves

Lawsonia inermis is widely known as henna and belongs to Lythraceae family. It contains various phytochemica constituents in every part of plant and shows various pharmacological properties. Secondary metabolites are acts as antioxidants and these are very high in this plant species. In this research studies, ethanolic extract of Lawsonia inermis was performed in the blood sample culture for cytotoxicity and genotoxicity effects. The both of the effects of plant extracts were evaluated through measurement of mitotic index (MI), sister chromatid exchanges (SCE) and cell proliferation kinetics (CPK). The blood sample culture was treated with control and various concentrations of extracts (500μg/ml and 1000μg/ml). The mean of MI, SCE and CPK for control were found to be 4.51, 5.19 and 0.52 respectively where as for 500μg/ml of Lawsonia inermis extracts are 4.33, 4.87 and 0.50 respectively and for 1000 μg/ml extracts are 4.18, 4.68 and 0.48 respectively. From the results, we can conclude that the MI and frequency of SCES of ethanolic plant extracts are remarkably similar to that of control which indicates that the Lawsonia inermis plant extracts have no substantial cytotoxicity and genotoxicity effects in cultured blood lymphocytes.

Marine toxin domoic acid induces moderate toxicological response in non-target HepG2 cells

Domoic acid (DA) is a marine neurotoxin produced as a defence compound by diatom Pseudo-nitzschia. Although its toxicity is well known in marine mammals and fish, data on DA cyto/genotoxicity in human non-target cells is still limited. Hence, we aimed to study the effect of DA (0.001–10 µg/mL) on cell viability and proliferation kinetics of human hepatocellular carcinoma (HepG2) cells as well as DNA damage induction after 4, 24 and 72 h of exposure. The results revealed that DA up to 10 µg/mL did not elicit significant changes in HepG2 cell viability, proliferation and cell cycle at applied conditions. DA did not generate DNA double-strand breaks, while it exhibited significant dose- and time-dependent increase of DNA damage in the form of either DNA single-strand breaks or alkali labile sites. Additionally, increased malondialdehyde level after DA treatment indicated oxidative damage to lipids. Altogether, the results showed that neurotoxin DA induced only minor adverse genotoxic effects in non-target HepG2 cells that most probably occurred resulting from the oxidative stress. However, additional research is needed to further elucidate the mechanisms of DA toxicity, particularly in terms of chronic exposure, as well as to understand its potential influence on human non-target cells.

A perspective on cell proliferation kinetics in the root apical meristem.

Organogenesis in plants is primarily postembryonic and relies on a strict balance between cell division and cell expansion. The root is a particularly well-suited model to study cell proliferation in detail since the two processes are spatially and temporally separated for all the different tissues. In addition, the root is amenable to detailed microscopic analysis to identify cells progressing through the cell cycle. While it is clear that cell proliferation activity is restricted to the root apical meristem (RAM), understanding cell proliferation kinetics and identifying its parameters have required much effort over many years. Here, we review the main concepts, experimental settings, and findings aimed at obtaining a detailed knowledge of how cells proliferate within the RAM. The combination of novel tools, experimental strategies, and mathematical models has contributed to our current view of cell proliferation in the RAM. We also discuss several lines of research that need to be explored in the future.

Real-Time Wireless Monitoring of Cell Proliferation and Detachment Based on pH-Responsive Conductive Polymer Dots.

In situ wireless monitoring for cell proliferation and detachment kinetics was conducted using pH-responsive zwitterionic polymer dots (Z-PDs), based on changes in electrochemical signals derived from Z-PD-coated substrates via the interaction of charges transferred between Z-PDs and cells. Z-PD-coated substrates were found to be a potent means to monitor and manipulate cell adhesion and detachment because of their high sensitivity over a wide range of pH conditions, and modification of the coated substrates was confirmed using a wireless system. At neutral pH, Z-PD-coated wireless sensors exhibited π-π stacking involving aromatic rings with hydrophobic interactions, thereby promoting cell proliferation; consequently, an increase in the measured resistance was observed. In contrast, Z-PD-coated substrates triggered by acidic and basic conditions promoted cell detachment, which induced an increase in the resistance compared with Z-PD substrates at pH 6.8, as a result of charges transferred to support Z-PD internalization through cell membranes after detachment. Therefore, as a wireless biosensor with excellent pH responsiveness that facilitates cell proliferation and detachment and whose electrochemical signals could be additionally acquired via a smartphone, Z-PD biosensors demonstrated a more favorable approach for monitoring cell-surface interactions than conventional optically based methods.

Mitochondrial activities play a pivotal role in regulating cell cycle in response to doxorubicin

ABSTRACT Doxorubicin induces both DNA damage and metabolic interference. How these effects interact to modulate cellular toxicity is not completely understood but important given the widespread use of doxorubicin in cancer treatment. This study tests the hypothesis that cell cycle arrest and survival are affected by distinct mitochondrial activities during doxorubicin exposure. Parental and mutant S. cerevisiae strains deficient in selected genes with mitochondrial function were treated with doxorubicin and assayed for changes in proliferation rates, cell survival and cell cycle arrest kinetics. Mitochondrial DNA content was estimated using quantitative PCR. Mitochondrial function was assessed by measuring oxygen consumption with and without an uncoupler. Parental cells growing in a non-fermentable carbon source medium and mutants lacking mitochondria and grown in glucose medium both show abrupt cell cycle and proliferation arrest during doxorubicin exposure compared to parental cells grown in glucose. Mitochondrial DNA increases during doxorubicin exposure in S. cerevisiae and in human breast cancer cells. Yeast strains deficient in TCA cycle activity or electron transport both show more abrupt cell cycle arrest than parental cells when exposed to doxorubicin. Concurrent treatment with the mitochondrial uncoupler dinitrophenol facilitates cell cycle progression and proliferation during doxorubicin exposure. Doxorubicin exposure induces mitochondrial DNA synthesis with TCA cycle and oxidative phosphorylation activity having opposing effects on cell proliferation, survival and cell cycle kinetics. TCA cycle activity provides biosynthetic substrates to support cell cycle progression and cell proliferation while electron transport and oxidative phosphorylation facilitate cell cycle arrest and possibly increased cytotoxicity.

Establishment of a Mouse Ovarian Cancer and Peritoneal Metastasis Model to Study Intraperitoneal Chemotherapy

Simple SummaryIntraperitoneal chemotherapy (IPC) is a locoregional treatment option in patients with peritoneal metastases (PM). Small animal models are valuable research tools allowing for rapid, reproducible, and inexpensive study and optimization of different forms of IPC. Here, we present a mouse model of ovarian cancer-derived PM and demonstrate its suitability for various modes of IPC, including pressurized intraperitoneal chemotherapy (PIPAC) using a micro-nozzle for aerosolized drug delivery.Intraperitoneal chemotherapy (IPC) is a locoregional treatment option in patients with peritoneal metastases (PM). Here, we present an ovarian cancer (OC)-derived PM mouse model for the study of different forms of IPC. Xenograft cell proliferation (luciferase-transfected OVCAR3 and SKOV3 clones) and growth kinetics were assessed using PET scan, bioluminescence imaging (BLI), and histological tumor analysis. Liquid IPC was achieved by intraperitoneal injection with/without capnoperitoneum (6–7 mmHg). Pressurized intraperitoneal aerosol chemotherapy (PIPAC) was mimicked using an intratracheal drug aerosol administration system (micro-nozzle), which, as demonstrated by ex vivo granulometric analysis using laser diffraction spectrometry, produced a polydisperse, bimodal aerosol with a volume-weighted median diameter of (26.49 ± 2.76) µm. Distribution of Tc-99m-labeled doxorubicin in mice was characterized using SPECT and was dependent on the delivery mode and most homogeneous when the micro-nozzle was used. A total of 2 mg doxorubicin per kg body weight was determined to be the optimally effective and tolerable dose to achieve at least 50% tumor reduction. Repeated PIPAC (four times at seven-day-intervals) with doxorubicin in SKOV3-luc tumor-bearing mice resulted in halted tumor proliferation and tumor load reduced after the second round of PIPAC versus controls and the number of tumor nodules was significantly reduced (27.7 ± 9.5 vs. 57.3 ± 9.5; p = 0.0003). Thus, we established the first mouse model of OC PM for the study of IPC using a human xenograft with SKOV3 cells and an experimental IPC setup with a miniaturized nozzle. Repeated IPC was feasible and demonstrated time-dependent anti-tumor activity.

An Experimental and Computational Protocol to Study Cell Proliferation in Human Acute Myeloid Leukemia Xenografts.

Acute myeloid leukemia (AML) is a highly frequent hematological malignancy, characterized by clinical and biological diversity, along with high relapse and mortality rates. The inherent functional and genetic intra-tumor heterogeneity in AML is thought to play an important role in disease recurrence and resistance to chemotherapy. Patient-derived xenograft (PDX) models preserve important features of the original tumor, allowing, at the same time, experimental manipulation and in vivo amplification of the human cells. Here we present a detailed protocol for the generation of fluorescently labeled AML PDX models to monitor cell proliferation kinetics in vivo, at the single-cell level. Although experimental protocols for cell proliferation studies are well established and widespread, they are not easily applicable to in vivo contexts, and the analysis of related time-series data is often complex to achieve. To overcome these limitations, model-driven approaches can be exploited to investigate different aspects of cell population dynamics. Among the existing approaches, the ProCell framework is able to perform detailed and accurate stochastic simulations of cell proliferation, relying on flow cytometry data. In particular, by providing an initial and a target fluorescence histogram, ProCell automatically assesses the validity of any user-defined scenario of intra-tumor heterogeneity, that is, it is able to infer the proportion of various cell subpopulations (including quiescent cells) and the division interval of proliferating cells. Here we explain the protocol in detail, providing a description of our methodology for the conditional expression of H2B-GFP in human AML xenografts, data processing by flow cytometry, and the final elaboration in ProCell.

Heterogeneous Polymeric Particles Encapsulating Human T cells for Controlled Activation, Proliferation, and Delivery.

We report a particulate cell delivery platform, toroidal spiral particles (TSPs), for continuous cell activation, expansion, and local sustained release. Biocompatible TSPs, generated by a self-assembly process of polymeric droplet sedimentation in an aqueous solution and subsequent polymer solidification, possess many engineering design flexibilities to manipulate the microenvironment of the cells to control cell proliferation, migration, and release kinetics. These millimeter-size particles with desired mechanical and physicochemical properties may be potentially used for adoptive cellular therapy (ACT) delivery by a minimally invasive procedure to the tumor mass.

Towards better combination regimens of cytarabine and FLT3 inhibitors in acute myeloid leukemia.

AML patients with FLT3/ITD mutations have poor response to cytarabine-based chemotherapy. FLT3 inhibitors (FLT3i) may resensitize cells to cytarabine (CYT). Improving treatment outcome of this combination may benefit from a mechanistic extrapolation approach from in vitro data. The effects of CYT and several FLT3i on cell proliferation and cell cycle kinetics were examined in AML cell lines. The effect of FLT3i (quizartinib, midostaurin, sorafenib) on cell proliferation and cell cycle kinetics was assessed in AML cell lines with differing FLT3 status; HEL (negligible expression of wild-type FLT3), EOL1 (wild-type FLT3), MV4-11 (FLT3-ITD resulting in constitutively active isoform). Semi-mechanistic cell cycle models for CYT and FLT3i were developed. Clinical CYT and quizartinib pharmacokinetic dosage regimens were modeled. Survival of AML patients was described via a hazard model. Simulations exploring different CYT/quizartinib regimens were conducted with the goal of improving treatment outcome. FLT3 status was associated with sensitivity to CYT (HEL cells most sensitive > EOL1 > MV4-11 cells). This order of sensitivity is reversed for FLT3i. Cytarabine induced apoptosis in the S-phase while all FLT3i induced apoptosis and cell cycle arrest at G1 phase. Simulations of candidate clinical regimens predict better cell kill upon adding quizartinib simultaneously with or immediately after CYT exposure. Overall survival was predicted to be significantly better with quizartinib 200mg administered every 48h vs every 24h in patients with FLT3 aberrations. Simultaneous administration of quizartinib and CYT every other day is a promising combination regimen for AML patients with FLT3 mutations.