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Endothelial dysfunction, prevalent in cardiovascular diseases (CVDs) and linked to conditions like diabetes, hypertension, obesity, renal failure, or hypercholesterolemia, is characterized by diminished nitric oxide (NO) bioavailability-a key signaling molecule for vascular homeostasis. Current two-dimensional (2D) in vitro studies on NO synthesis by endothelial cells (ECs) lack the crucial laminar shear stress, a vital factor in modulating the NO-generating enzyme, endothelial nitric oxide synthase (eNOS), under physiological conditions. Here we developed a tracer-based metabolomics approach to measure NO-specific metabolites with mass spectrometry (MS) and show the impact of fluid flow on metabolic parameters associated with NO synthesis using 2D and 3D platforms. Specifically, we tracked the conversion of stable-isotope labeled NO substrate L-Arginine to L-Citrulline and L-Ornithine to determine eNOS activity. We demonstrated clear responses in human coronary artery endothelial cells (HCAECs) cultured with 13C6, 15N4-L-Arginine, and treated with eNOS stimulator, eNOS inhibitor, and arginase inhibitor. Analysis of downstream metabolites, 13C6, 15N3 L-Citrulline and 13C5, 15N2 L-Ornithine, revealed distinct outcomes. Additionally, we evaluated the NO metabolic status in static 2D culture and 3D microvessel models with bidirectional and unidirectional fluid flow. Our 3D model exhibited significant effects, particularly in microvessels exposed to the eNOS stimulator, as indicated by the 13C6, 15N3 L-Citrulline/13C5, 15N2 L-Ornithine ratio, compared to the 2D culture. The obtained results indicate that the 2D static culture mimics an endothelial dysfunction status, while the 3D model with a unidirectional fluid flow provides a more representative physiological environment that provides a better model to study endothelial dysfunction.
Subject(s)
Endothelial Cells , Metabolomics , Microvessels , Nitric Oxide Synthase Type III , Nitric Oxide , Humans , Nitric Oxide/metabolism , Metabolomics/methods , Microvessels/metabolism , Nitric Oxide Synthase Type III/metabolism , Endothelial Cells/metabolism , Arginine/metabolism , Lab-On-A-Chip Devices , Cells, Cultured , Citrulline/metabolismABSTRACT
BACKGROUND: Abnormal uterine bleeding (AUB) that occurs in a structurally normal uterus with regular menstrual cycles and without other identifiable etiology is often caused by a primary endometrial disorder (AUB-E). Altered vascular morphological changes and expression of markers of angiogenesis have been implicated as an underlying cause in these cases. OBJECTIVES: The study was conducted to investigate the expression of vascular endothelial growth factor (VEGF) and smooth muscle actin-alpha (SMA-α), and to perform microvessel density (MVD), and morphometric evaluation of endometrial vessels in patients with AUB-E. MATERIAL AND METHODS: Endometrial biopsies and hysterectomy specimens of 40 patients clinically diagnosed with AUB-E were included in the study with 40 age-matched controls. Immunohistochemistry (IHC) with VEGF and SMA-α was performed, and the expression and staining pattern was recorded as the number of positive vessels per 10 high power fields and intensity scores. Morphometric analysis was performed on CD34 stained sections using Leica Application Suite, version 4.4.0 software (Leica Microsystems, Wetzlar, Germany). MVD was calculated by the vascular hotspot method. RESULTS: A statistically significant increase in VEGF vessel count (p-value<0.001) and a decline in SMA-α expression (p-value=0.23) was seen in cases as compared to the control group. There was a statistically significant increase in microvessel caliber (p-value=0.01) and MVD (p-value <0.001) in cases as compared to controls. CONCLUSION: These findings support aberrant vascular proliferation and impaired vessel maturation, contributing to the pathology of AUB-E. Alterations in angiogenesis in these patients reveal potential therapeutic targets for AUB.
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Background and Purpose: Impaired cerebral circulation, induced by blood vessel constrictions and microthrombi, leads to delayed cerebral ischemia after subarachnoid hemorrhage (SAH). 12/15-Lipooxygenase (12/15-LOX) overexpression has been implicated in worsening early brain injury outcomes following SAH. However, it is unknown if 12/15-LOX is important in delayed pathophysiological events after SAH. Since 12/15-LOX produces metabolites that induce inflammation and vasoconstriction, we hypothesized that 12/15-LOX leads to microvessel constriction and microthrombi formation after SAH, and thus 12/15-LOX is an important target to prevent delayed cerebral ischemia. Methods: SAH was induced in C57BL/6 and 12/15-LOX-/- mice of both sexes by endovascular perforation. Expression of 12/15-LOX was assessed in brain tissue slices and in vitro. C57BL/6 mice were administered either ML351 (12/15-LOX inhibitor) or vehicle. Mice were evaluated for daily neuroscore and euthanized on day five to assess cerebral 12/15-LOX expression, vessel constrictions, platelet activation, microthrombi, neurodegeneration, infarction, cortical perfusion, and for development of delayed deficits. Finally, the effect of 12/15-LOX inhibition on platelet activation was assessed in SAH patient samples using a platelet spreading assay. Results: In SAH mice, 12/15-LOX was upregulated in brain vascular cells and there was an increase in 12-S-HETE. Inhibition of 12/15-LOX improved brain perfusion on days 4-5 and attenuated delayed pathophysiological events, including microvessel constrictions, microthrombi, neuronal degeneration, and infarction. Additionally, 12/15-LOX inhibition reduced platelet activation in human and mouse blood samples. Conclusions: Cerebrovascular 12/15-LOX overexpression plays a major role in brain dysfunction after SAH by triggering microvessel constrictions and microthrombi formation, which reduces brain perfusion. Inhibiting 12/15-LOX may be a therapeutic target to improve outcomes after SAH.
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The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), responsible for the COVID-19, may lead to multiple organ dysfunctions and long-term complications. The induction of microvascular dysfunction is regarded as a main player in these pathological processes. To investigate the possible impact of SARS-CoV-2-induced endothelial-to-mesenchymal transition (EndMT) on fibrosis in "long-COVID" syndrome, we used primary cultures of human microvascular cells derived from the lungs, as the main infection target, compared to cells derived from different organs (dermis, heart, kidney, liver, brain) and to the HUVEC cell line. To mimic the virus action, we used mixed SARS-CoV-2 peptide fragments (PepTivator®) of spike (S), nucleocapsid (N), and membrane (M) proteins. TGFß2 and cytokine mix (IL-1ß, IL-6, TNFα) were used as positive controls. The percentage of cells positive to mesenchymal and endothelial markers was quantified by high content screening. We demonstrated that S+N+M mix induces irreversible EndMT in all analyzed endothelial cells via the TGFß pathway, as demonstrated by ApoA1 treatment. We then tested the contribution of single peptides in lung and brain cells, demonstrating that EndMT is triggered by M peptide. This was confirmed by transfection experiment, inducing the endogenous expression of the glycoprotein M in lung-derived cells. In conclusion, we demonstrated that SARS-CoV-2 peptides induce EndMT in microvascular endothelial cells from multiple body districts. The different peptides play different roles in the induction and maintenance of the virus-mediated effects, which are organ-specific. These results corroborate the hypothesis of the SARS-CoV-2-mediated microvascular damage underlying the multiple organ dysfunctions and the long-COVID syndrome.
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Intrahepatic cholangiocarcinoma (iCCA) exhibits different blood imaging features and prognosis depending on histology. To clarity histopathological growth patterns (HGPs) and vascularization processes of iCCA, we collected 145 surgical specimens and histologically classified them into large bile duct (LBD) (20 cases), small bile duct (SBD) (54), cholangiolocarcinoma (CLC) (35), combined SBD-CLC (cSBD-CLC) (26), and ductal plate malformation (DPM) (10) (sub)types. According to the invasive pattern at the interface between tumor and adjacent background liver, HGPs were classified into desmoplastic, pushing, and replacing HGPs. Desmoplastic HGP predominated in LBD type (55.5%), while replacing HGP was common in CLC (82.9%) and cSBD-CLC (84.6%) subtypes. Desmoplastic HGP reflected angiogenesis, while replacing HGP showed vessel co-option in addition to angiogenesis. By evaluating microvessel density (MVD) using vascular markers, ELTD1 identified vessel co-option and angiogenesis, and ELTD1-positive MVD at invasive margin in replacing HGP was significantly higher than those in desmoplastic and pushing HGPs. REDD1, an angiogenesis-related marker, demonstrated preferably higher MVD in the tumor center than in other areas. iCCA (sub)types and HGPs were closely related to vessel co-option and immune-related factors (lymphatic vessels, lymphocytes, and neutrophils). In conclusion, HGPs and vascular mechanisms characterize iCCA (sub)types and vessel co-option linked to the immune microenvironment.
Subject(s)
Bile Duct Neoplasms , Cholangiocarcinoma , Neovascularization, Pathologic , Cholangiocarcinoma/pathology , Cholangiocarcinoma/blood supply , Humans , Bile Duct Neoplasms/pathology , Male , Female , Neovascularization, Pathologic/pathology , Middle Aged , Aged , Adult , Microvascular DensityABSTRACT
Single-agent TAS102 (trifluridine/tipiracil) and regorafenib are FDA-approved treatments for metastatic colorectal cancer (mCRC). We previously reported that regorafenib combined with a fluoropyrimidine can delay disease progression in clinical case reports of multidrug-resistant mCRC patients. We hypothesized that the combination of TAS102 and regorafenib may be active in CRC and other gastrointestinal (GI) cancers and may in the future provide a treatment option for patients with advanced GI cancer. We investigated the therapeutic effect of TAS102 in combination with regorafenib in preclinical studies employing cell culture, colonosphere assays that enrich for cancer stem cells, and in vivo. TAS102 in combination with regorafenib has synergistic activity against multiple GI cancers in vitro including colorectal and gastric cancer, but not liver cancer cells. TAS102 inhibits colonosphere formation and this effect is potentiated by regorafenib. In vivo anti-tumor effects of TAS102 plus regorafenib appear to be due to anti-proliferative effects, necrosis and angiogenesis inhibition. Growth inhibition by TAS102 plus regorafenib occurs in xenografted tumors regardless of p53, KRAS or BRAF mutations, although more potent tumor suppression was observed with wild-type p53. Regorafenib significantly inhibits TAS102-induced angiogenesis and microvessel density in xenografted tumors, as well inhibits TAS102-induced ERK1/2 activation regardless of RAS or BRAF status in vivo. TAS102 plus regorafenib is a synergistic drug combination in preclinical models of GI cancer, with regorafenib suppressing TAS102-induced increase in microvessel density and p-ERK as contributing mechanisms. The TAS102 plus regorafenib drug combination may be further tested in gastric and other GI cancers.
Subject(s)
Drug Combinations , Drug Synergism , Gastrointestinal Neoplasms , Mutation , Neoplastic Stem Cells , Neovascularization, Pathologic , Phenylurea Compounds , Proto-Oncogene Proteins B-raf , Proto-Oncogene Proteins p21(ras) , Pyridines , Pyrrolidines , STAT3 Transcription Factor , Thymine , Trifluridine , Uracil , Xenograft Model Antitumor Assays , Humans , Trifluridine/pharmacology , Phenylurea Compounds/pharmacology , Animals , Pyridines/pharmacology , Proto-Oncogene Proteins B-raf/genetics , Proto-Oncogene Proteins B-raf/metabolism , Proto-Oncogene Proteins p21(ras)/genetics , Proto-Oncogene Proteins p21(ras)/metabolism , Neovascularization, Pathologic/drug therapy , Neovascularization, Pathologic/genetics , Neovascularization, Pathologic/metabolism , Neoplastic Stem Cells/drug effects , Neoplastic Stem Cells/metabolism , Neoplastic Stem Cells/pathology , Gastrointestinal Neoplasms/drug therapy , Gastrointestinal Neoplasms/genetics , Gastrointestinal Neoplasms/pathology , Gastrointestinal Neoplasms/metabolism , Uracil/pharmacology , Uracil/analogs & derivatives , Mice , STAT3 Transcription Factor/metabolism , STAT3 Transcription Factor/genetics , Thymine/pharmacology , Cell Line, Tumor , Pyrrolidines/pharmacology , Pyrrolidines/therapeutic use , Antineoplastic Combined Chemotherapy Protocols/pharmacology , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , MAP Kinase Signaling System/drug effects , Signal Transduction/drug effects , Cell Proliferation/drug effects , AngiogenesisABSTRACT
OBJECTIVE: Tortuous microvessels are characteristic of microvascular remodeling associated with numerous physiological and pathological scenarios. Three-dimensional (3D) hemodynamics in tortuous microvessels influenced by red blood cells (RBCs), however, are largely unknown, and important questions remain. Is blood viscosity influenced by vessel tortuosity? How do RBC dynamics affect wall shear stress (WSS) patterns and the near-wall cell-free layer (CFL) over a range of conditions? The objective of this work was to parameterize hemodynamic characteristics unique to a tortuous microvessel. METHODS: RBC-resolved simulations were performed using an immersed boundary method-based 3D fluid dynamics solver. A representative tortuous microvessel was selected from a stimulated angiogenic network obtained from imaging of the rat mesentery and digitally reconstructed for the simulations. The representative microvessel was a venule with a diameter of approximately 20 µm. The model assumes a constant diameter along the vessel length and does not consider variations due to endothelial cell shapes or the endothelial surface layer. RESULTS: Microvessel tortuosity was observed to increase blood apparent viscosity compared to a straight tube by up to 26%. WSS spatial variations in high curvature regions reached 23.6 dyne/cm2 over the vessel cross-section. The magnitudes of WSS and CFL thickness variations due to tortuosity were strongly influenced by shear rate and negligibly influenced by tube hematocrit levels. CONCLUSIONS: New findings from this work reveal unique tortuosity-dependent hemodynamic characteristics over a range of conditions. The results provide new thought-provoking information to better understand the contribution of tortuous vessels in physiological and pathological processes and help improve reduced-order models.
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Introduction: The dynamics of circulating tumor cells (CTCs) within blood vessels play a pivotal role in predicting metastatic spreading of cancer within the body. However, the limited understanding and method to quantitatively investigate the influence of vascular architecture on CTC dynamics hinders our ability to predict metastatic process effectively. To address this limitation, the present study was conducted to investigate the influence of blood vessel tortuosity on the behaviour of CTCs, focusing specifically on establishing methods and examining the role of shear stress in CTC-vessel wall interactions and its subsequent impact on metastasis. Methods: We computationally simulated CTC behaviour under various shear stress conditions induced by vessel tortuosity. Our computational model, based on the lattice Boltzmann method (LBM) and a coarse-grained spectrin-link membrane model, efficiently simulates blood plasma dynamics and CTC deformability. The model incorporates fluid-structure interactions and receptor-ligand interactions crucial for CTC adhesion using the immersed boundary method (IBM). Results: Our findings reveal that uniform shear stress in straight vessels leads to predictable CTC-vessel interactions, whereas in curved vessels, asymmetrical flow patterns and altered shear stress create distinct adhesion dynamics, potentially influencing CTC extravasation. Quantitative analysis shows a 25% decrease in the wall shear stress in low-shear regions and a 58.5% increase in the high-shear region. We observed high-shear regions in curved vessels to be potential sites for increased CTC adhesion and extravasation, facilitated by elevated endothelial expression of adhesion molecules. This phenomenon correlates with the increased number of adhesion bonds, which rises to approximately 40 in high-shear regions, compared to around 12 for straight vessels and approximately 5-6 in low-shear regions. The findings also indicate an optimal cellular stiffness necessary for successful CTC extravasation in curved vessels. Discussion: By the quantitative assessment of the risk of CTC extravasation as a function of vessel tortuosity, our study offers a novel tool for the prediction of metastasis risk to support the development of personalized therapeutic interventions based on individual vascular characteristics and tumor cell properties.
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Vasculature-on-chip (VoC) models have become a prominent tool in the study of microvasculature functions because of their cost-effective and ethical production process. These models typically use a hydrogel in which the three-dimensional (3D) microvascular structure is embedded. Thus, VoCs are directly impacted by the physical and chemical cues of the supporting hydrogel. Endothelial cell (EC) response in VoCs is critical, especially in organ-specific vasculature models, in which ECs exhibit specific traits and behaviors that vary between organs. Many studies customize the stimuli ECs perceive in different ways; however, customizing the hydrogel composition accordingly to the target organ's extracellular matrix (ECM), which we believe has great potential, has been rarely investigated. We explored this approach to organ-specific VoCs by fabricating microvessels (MVs) with either human umbilical vein ECs or human brain microvascular ECs in a 3D cylindrical VoC using a collagen hydrogel alone or one supplemented with laminin and hyaluronan, components found in the brain ECM. We characterized the physical properties of these hydrogels and analyzed the barrier properties of the MVs. Barrier function and tight junction (ZO-1) expression improved with the addition of laminin and hyaluronan in the composite hydrogel.
Subject(s)
Collagen , Human Umbilical Vein Endothelial Cells , Hyaluronic Acid , Hydrogels , Laminin , Microvessels , Tight Junctions , Humans , Hydrogels/chemistry , Hyaluronic Acid/chemistry , Hyaluronic Acid/pharmacology , Laminin/chemistry , Laminin/metabolism , Collagen/chemistry , Collagen/metabolism , Microvessels/metabolism , Microvessels/drug effects , Tight Junctions/metabolism , Human Umbilical Vein Endothelial Cells/metabolism , Lab-On-A-Chip Devices , Endothelial Cells/metabolism , Endothelial Cells/drug effects , Cells, CulturedABSTRACT
A hallmark of cerebral malaria is sequestration of Plasmodium falciparum-infected erythrocytes (IEs) in the brain microcirculation. Antibodies contribute to malaria immunity, but it remains unclear whether functional antibodies targeting parasite-expressed ligand can block cytoadhesion in the brain. Here, we screened the plasma of older children and young adults in Malawi to characterize the antibody response against the P. falciparum-IE surface and used a bioengineered 3D human brain microvessel model incorporating variable flow dynamics to measure adhesion blocking responses. We found a strong correlation between surface antibody reactivity by flow cytometry and reduced P. falciparum-IE binding in 3D microvessels. Moreover, there was a threshold of surface antibody reactivity necessary to achieve robust inhibitory activity. Our findings provide evidence of the acquisition of adhesion blocking antibodies against cerebral binding variants in people exposed to stable P. falciparum transmission and suggest the quality of the inhibitory response can be influenced by flow dynamics.
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Red blood cells (RBCs) carry oxygen and make up 40-45% of blood by volume in large vessels down to 10% or less in smaller capillaries. Because of their finite size and large volume fraction, they are heterogeneously distributed throughout the body. This is partially because RBCs are distributed or partitioned nonuniformly at diverging vessel bifurcations where blood flows from one vessel into two. Despite its increased recognition as an important player in the microvasculature, few studies have explored how the endothelial surface layer (ESL; a vessel wall coating) may affect partitioning and RBC dynamics at diverging vessel bifurcations. Here, we use a mathematical and computational model to consider how altering ESL properties, as can occur in pathological scenarios, change RBC partitioning, deformation, and penetration of the ESL. The two-dimensional finite element model considers pairs of cells, represented by interconnected viscoelastic elements, passing through an ESL-lined diverging vessel bifurcation. The properties of the ESL include the hydraulic resistivity and an osmotic pressure difference modeling how easily fluid flows through the ESL and how easily the ESL is structurally compressed, respectively. We find that cell-cell interaction leads to more uniform partitioning and greatly enhances the effects of ESL properties, especially for deformation and penetration. This includes the trend that increased hydraulic resistivity leads to more uniform partitioning, increased deformation, and decreased penetration. It also includes the trend that decreased osmotic pressure increases penetration.
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Blood flow in arterioles have attracted considerable research attention due to their clinical implications. However, the fluid structure interaction between red blood cells and plasma in the blood poses formidable difficulty to the computational efforts. In this contribution, we seek to represent the red blood cells in the blood as a continuous non-Newtonian phase and construct a multi-phase model for the blood flow in microvessels. The methods are presented and validated using a channel with sudden expansion. And the resulting blood flow inside a stenosed microvessel is investigated at different inlet velocity amplitudes and hematocrits. It is show that the increase of both inlet velocity amplitude and inlet hematocrit leads to longer and thicker cell-rich layer downstream the stenosis. Besides, it is found that the maximum values of wall shear stress scales up with inlet velocity amplitudes and hematocrits. These results show the validity of the proposed computational model and provide helpful insights into blood flow behaviors inside stenosed vessels.
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OBJECTIVE: Evaluation of the predictive value of one-stop energy spectrum and perfusion CT parameters for microvessel density (MVD) in colorectal cancer cancer foci. METHODS: Clinical and CT data of 82 patients with colorectal cancer confirmed by preoperative colonoscopy or surgical pathology in our hospital from September 2019 to November 2022 were collected and analyzed retrospectively. Energy spectrum CT images were measured using the Protocols general module of the GSI Viewer software of the GE AW 4.7 post-processing workstation to measure the CT values of the arterial and venous phase lesions and the neighboring normal intestinal wall in a single energy range of 40 kevâ¼140 kev, and the slopes of the energy spectrum curves (λ) were calculated between 40 kev-90 kev; Iodine concentration (IC), Water concentration (WC), Effective-Z (Eff-Z) and Normalized iodine concentration (NIC) were measured by placing a region of interest (ROI) on the iodine concentration map and water concentration map at the lesion and adjacent to the normal intestinal wall.Perfusion CT images were scanned continuously and dynamically using GSI Perfusion software and analyzed by applying CT Perfusion 4.0 software.Blood volume (BV), blood flow (BF), surface permeability (PS), time to peak (TTP), and mean transit time (MTT) were measured respectively in the lesion and adjacent normal colorectal wall. Based on the pathological findings, the tumors were divided into a low MVD group (MVD < 35/field of view, n = 52 cases) and a high MVD group (MVD ≥ 35/field of view, n = 30 cases) using a median of 35/field of view as the MVD grouping criterion. The collected data were statistically analyzed, the subjects' operating characteristic curve (ROC) was plotted, and the area under curve (AUC), sensitivity, specificity, and Yoden index were calculated for the predicted efficacy of each parameter of the energy spectrum and perfusion CT and the combined parameters. RESULTS: The CT values, IC, NIC, λ, Eff-Z of 40kevâ¼140kev single energy in the arterial and venous phase of colorectal cancer in the high MVD group were higher than those in the low MVD group, and the differences were all statistically significant (p < 0.05). The AUC of each single-energy CT value in the arterial phase from 40 kev to 120 kev for determining the high or low MVD of colorectal cancer was greater than 0.8, indicating that arterial stage has a good predictive value for high or low MVD in colorectal cancer; AUC for arterial IC, NIC and IC + NIC were all greater than 0.9, indicating that in arterial colorectal cancer, both single and combined parameters of spectral CT are highly effective in predicting the level of MVD. The AUC of 40 kev to 90 kev single-energy CT values in the intravenous phase was greater than 0.9, and its diagnostic efficacy was more representative; The AUC of IC and NIC in venous stage were greater than 0.8, which indicating that the IC and NIC energy spectrum parameters in venous stage colorectal cancer have a very good predictive value for the difference between high and low MVDs, with the greatest diagnostic efficacy in IC.The values of BV and BF in the high MVD group were higher than those in the low MVD group, and the differences were statistically significant (P < 0.05), and the AUC of BF, BV, and BV + BF were 0.991, 0.733, and 0.997, respectively, with the highest diagnostic efficacy for determining the level of MVD in colorectal cancer by BV + BF. CONCLUSION: One-stop CT energy spectrum and perfusion imaging technology can accurately reflect the MVD in living tumor tissues, which in turn reflects the tumor angiogenesis, and to a certain extent helps to determine the malignancy, invasion and metastasis of living colorectal cancer tumor tissues based on CT energy spectrum and perfusion parameters.
Subject(s)
Neovascularization, Pathologic , Humans , Male , Female , Middle Aged , Aged , Neovascularization, Pathologic/diagnostic imaging , Retrospective Studies , Tomography, X-Ray Computed/methods , Adult , Rectal Neoplasms/diagnostic imaging , Rectal Neoplasms/blood supply , Rectal Neoplasms/pathology , Aged, 80 and over , Microvascular Density , Colorectal Neoplasms/diagnostic imaging , Colorectal Neoplasms/blood supply , Colorectal Neoplasms/pathology , Predictive Value of Tests , Colonic Neoplasms/diagnostic imaging , Colonic Neoplasms/blood supply , AngiogenesisABSTRACT
Angiogenesis is considered essential for tumor progression; however, whether histological counting of blood vessel numbers, expressed as microvessel density (MVD), can be a prognostic factor in breast cancer remains controversial. It has been suggested that the specific morphology of blood vessels such as glomeruloid microvascular proliferation (GMP) is associated with clinical parameters. Here, we aimed to clarify the significance of MVD with revised immunohistochemistry and to identify new blood vessel shapes that predict prognosis in breast cancer. Four hundred and eleven primary breast cancer specimens were collected, and the sections were immunohistochemically stained with CD31 (single staining) and CD31 and Collagen IV (double staining). The prognosis of patients was examined based on the MVD value, and the presence of GMP and other blood vessels with other specific shapes. As a result, high MVD value and the presence of GMP were not associated with worse prognosis. By contrast, patients with deep-curved capillaries surrounding tumor cell nests (C-shaped) or excessively branched capillaries near tumor cell nests showed a significantly poor prognosis. The presence of these capillaries was also correlated with clinicopathological parameters such as Ki-67 index. Thus, the morphology of capillaries rather than MVD can be a better indicator of tumor aggressiveness.
Subject(s)
Breast Neoplasms , Capillaries , Microvascular Density , Neovascularization, Pathologic , Humans , Female , Breast Neoplasms/pathology , Breast Neoplasms/blood supply , Prognosis , Middle Aged , Neovascularization, Pathologic/pathology , Aged , Adult , Capillaries/pathology , Immunohistochemistry , Aged, 80 and over , Biomarkers, Tumor/analysis , Biomarkers, Tumor/metabolismABSTRACT
Peroxiredoxins (Prxs) are a ubiquitously expressed family of antioxidant enzymes that either facilitate or inhibit tumorigenesis, depending on the cancer type and Prx isoform. Prx2 is a typical Prx that has a dual role in tumorigenesis and tumor progression. However, the expression of Prx2 and its precise role in cervical cancer remains to be elucidated. Therefore, the present study aimed to investigate the expression of Prx2 and its association with the progression and prognosis of cervical squamous cell cancer (CSCC). In the present study, the clinicopathological data of 105 patients diagnosed with CSCC were collected from the medical record system at Jingzhou Central Hospital, Tongji Medical College of Huazhong University of Science and Technology (Jingzhou, China). Prx2 protein was also detected in 105 CSCC tissues and 40 adjacent peri-tumoral tissues by immunohistochemical staining. The relationships between Prx2 expression and clinicopathological features, vascular endothelial growth factor A (VEGF-A) expression and micro-vessel density (MVD) in CSCC were then analyzed. Progression-free survival (PFS) was also assessed using both univariate and multivariate analyses. The results of the present study demonstrated that the expression of Prx2 was upregulated in CSCC tissues compared with the adjacent peri-tumoral tissues (P<0.001). In addition, higher Prx2 expression was associated with greater depth of stromal invasion (P=0.023) and positive lymph vascular space invasion (P=0.044), while the Prx2 expression level was not associated with age, tumor size, histological grade, lymph node (LN) metastasis or International Federation of Gynecology and Obstetrics (FIGO) stage (all P>0.05). Furthermore, increased Prx2 expression was associated with high MVD (P=0.016), while expression of VEGF-A was not associated with Prx2 expression (P>0.05). Kaplan-Meier analysis showed that patients with high Prx2 expression (log-rank test, P=0.039), high MVD (log-rank test, P=0.015), a higher FIGO stage (log-rank test, P=0.021) and LN metastasis (log-rank test, P=0.022) had a shorter PFS time than patients with low Prx2 expression, low MVD, a lower FIGO stage and without LN metastasis, respectively. Cox proportional hazard regression analysis revealed that expression of Prx2 [hazard ratio (HR), 2.551; 95% confidence interval (CI), 1.056-6.162; P=0.037], MVD (HR, 2.436; CI, 1.034-5.735; P=0.042) and FIGO stage (HR, 1.543; CI, 1.027-2.319; P=0.037) were independent factors for PFS time. In conclusion, the results of the present study suggested that Prx2 could act as a potential biomarker for predicting CSCC progression and prognosis and could be a novel target for antiangiogenic therapy of CSCC.
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BACKGROUND: The incidence and mortality of colorectal cancer (CRC) are among the highest in the world, and its occurrence and development are closely related to tumor neovascularization. When the balance between pigment epithelium-derived factors (PEDF) that inhibit angiogenesis and vascular endothelial growth factors (VEGF) that stimulate angiogenesis is broken, angiogenesis is out of control, resulting in tumor development. Therefore, it is very necessary to find more therapeutic targets for CRC for early intervention and later treatment. AIM: To investigate the expression and significance of PEDF, VEGF, and CD31-stained microvessel density values (CD31-MVD) in normal colorectal mucosa, adenoma, and CRC. METHODS: In this case-control study, we collected archived wax blocks of specimens from the Digestive Endoscopy Center and the General Surgery Department of Chengdu Second People's Hospital from April 2022 to October 2022. Fifty cases of specimen wax blocks were selected as normal intestinal mucosa confirmed by electronic colonoscopy and concurrent biopsy (normal control group), 50 cases of specimen wax blocks were selected as colorectal adenoma confirmed by electronic colonoscopy and pathological biopsy (adenoma group), and 50 cases of specimen wax blocks were selected as CRC confirmed by postoperative pathological biopsy after inpatient operation of general surgery (CRC group). An immunohistochemical staining experiment was carried out to detect PEDF and VEGF expression in three groups of specimens, analyze their differences, study the relationship between the two and clinicopathological factors in CRC group, record CD31-MVD in the three groups, and analyze the correlation of PEDF, VEGF, and CD31-MVD in the colorectal adenoma group and the CRC group. The F test or adjusted F test is used to analyze measurement data statistically. Kruskal-Wallis rank sum test was used between groups for ranked data. The chi-square test, adjusted chi-square test, or Fisher's exact test were used to compare the rates between groups. All differences between groups were compared using the Bonferroni method for multiple comparisons. Spearman correlation analysis was used to test the correlation of the data. The test level (α) was 0.05, and a two-sided P< 0.05 was considered statistically significant. RESULTS: The positive expression rate and expression intensity of PEDF were gradually decreased in the normal control group, adenoma group, and CRC group (100% vs 78% vs 50%, χ2 = 34.430, P < 0.001; ++~++ vs +~++ vs -~+, H = 94.059, P < 0.001), while VEGF increased gradually (0% vs 68% vs 96%, χ2 = 98.35, P < 0.001; - vs -~+ vs ++~+++, H = 107.734, P < 0.001). In the CRC group, the positive expression rate of PEDF decreased with the increase of differentiation degree, invasion depth, lymph node metastasis, distant metastasis, and TNM stage (χ2 = 20.513, 4.160, 5.128, 6.349, 5.128, P < 0.05); the high expression rate of VEGF was the opposite (χ2 = 10.317, 13.134, 17.643, 21.844, 17.643, P < 0.05). In the colorectal adenoma group, the expression intensity of PEDF correlated negatively with CD31-MVD (r = -0.601, P < 0.001), whereas VEGF was not significantly different (r = 0.258, P = 0.07). In the CRC group, the expression intensity of PEDF correlated negatively with the expression intensity of CD31-MVD and VEGF (r = -0.297, P < 0.05; r = -0.548, P < 0.05), while VEGF expression intensity was positively related to CD31-MVD (r = 0.421, P = 0.002). CONCLUSION: It is possible that PEDF can be used as a new treatment and prevention target for CRC by upregulating the expression of PEDF while inhibiting the expression of VEGF.
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The microvasculature is integral to nearly every tissue in the body, providing not only perfusion to and from the tissue, but also homing sites for immune cells, cellular niches for tissue dynamics, and cooperative interactions with other tissue elements. As a microtissue itself, the microvasculature is a composite of multiple cell types exquisitely organized into structures (individual vessel segments and extensive vessel networks) capable of considerable dynamics and plasticity. Consequently, it has been challenging to include a functional microvasculature in assembled or fabricated tissues. Isolated fragments of intact microvessels, which retain the cellular composition and structures of native microvessels, are proving effective in a variety of vascularization applications including tissue in vitro disease modeling, vascular biology, mechanistic discovery, and tissue prevascularization in regenerative therapeutics and grafting. In this review, we will discuss the importance of recapitulating native tissue biology and the successful vascularization applications of isolated microvessels.
Subject(s)
Microvessels , Neovascularization, Physiologic , Humans , Microvessels/physiology , AnimalsABSTRACT
Modeling organ-blood barriers through the inclusion of microvessel networks within in vitro tissue models could lead to more physiologically accurate results, especially since organ-blood barriers are crucial to the normal function, drug transport, and disease states of vascularized organs. Microvessel networks are difficult to form, since they push the practical limits of most fabrication methods, and it is difficult to coax vascular cells to self-assemble into structures larger than capillaries. Here, we present a method for rapidly forming networks of microvessel-like structures using sacrificial alginate structures. Specifically, we encapsulated endothelial cells within short alginate threads, and then embedded them in collagen gel. Following enzymatic degradation of the alginate, the collagen gel contained a network of hollow channels seeded with cells, all surrounding a perfusable central channel. This method uses a 3D-printed coaxial extruder and syringe pumps to generate short threads in a way that is repeatable and easily transferrable to other labs. The cell-laden, sacrificial alginate threads can be frozen after fabrication and thawed before embedding without significant loss of cell viability. The ability to freeze the threads enables future scale-up and ease of use. Within millifluidic devices that restrict access to media, the threads enhance cell survival under static conditions. These results indicate the potential for use of this method in a range of tissue engineering applications.
Subject(s)
Alginates , Microvessels , Tissue Engineering , Alginates/chemistry , Microvessels/cytology , Humans , Tissue Engineering/methods , Human Umbilical Vein Endothelial Cells/cytology , Human Umbilical Vein Endothelial Cells/metabolism , Tissue Scaffolds/chemistry , Endothelial Cells/cytology , Endothelial Cells/metabolism , Glucuronic Acid/chemistry , Hexuronic Acids/chemistry , Cell Survival , Animals , Collagen/chemistryABSTRACT
The use of [18F]FDG PET/CT as a biomarker in diffuse lung diseases is increasingly recognized. We investigated the correlation between [18F]FDG uptake with histologic markers on lung biopsy of patients with fibrotic interstitial lung disease (fILD). Methods: We recruited 18 patients with fILD awaiting lung biopsy for [18F]FDG PET/CT. We derived a target-to-background ratio (TBR) of maximum pulmonary uptake of [18F]FDG (SUVmax) divided by the lung background (SUVmin). Consecutive paraffin-embedded lung biopsy sections were immunostained for alveolar and interstitial macrophages (CD68), microvessel density (MVD) (CD31 and CD105/endoglin), and glucose transporter 1. MVD was expressed as vessel area percentage per high-power field (Va%/hpf). Differences in imaging and angiogenesis markers between histologic usual interstitial pneumonia (UIP) and non-UIP were assessed using a nonparametric Mann-Whitney test. Correlation of imaging with angiogenesis markers was assessed using the nonparametric Spearman rank correlation. Univariate Kaplan-Meier survival analysis assessed the difference in the survival curves for each of the angiogenesis markers (separated by their respective optimal cutoff) using the log-rank test. Statistical analysis was performed using SPSS. Results: In total, 18 patients were followed for an average of 41.36 mo (range, 5.69-132.46 mo; median, 30.07 mo). Only CD105 MVD showed a significantly positive correlation with [18F]FDG TBR (Spearman rank correlation, 0.556; P < 0.05, n = 13). There was no correlation between [18F]FDG uptake and macrophage expression of glucose transporter 1. CD105 and CD31 were higher for UIP than for non-UIP, with CD105 reaching statistical significance (P = 0.011). In all patients, MVD assessed with either CD105 or CD31 quantification on biopsy predicted overall survival. Patients with CD105 MVD of less than 12 Va%/hpf or CD31 MVD of less than 35 Va%/hpf had a significantly better prognosis (no deaths during follow-up in the case of CD105) than did patients with higher scores of CD105 MVD (median survival, 35 mo; P = 0.041, n = 13) or CD31 MVD (median survival, 28 mo; P = 0.014, n = 13). Conclusion: Previous work has used [18F]FDG uptake in PET/CT as a biomarker in fILD. Here, we highlight a correlation between angiogenesis and [18F]FDG TBR. We show that MVD is higher for UIP than for non-UIP and is associated with mortality in patients with fILD. These data set the scene to investigate the potential role of vasculature and angiogenesis in fibrosis.
Subject(s)
Lung Diseases, Interstitial , Positron Emission Tomography Computed Tomography , Humans , Positron Emission Tomography Computed Tomography/methods , Fluorodeoxyglucose F18 , Glucose Transporter Type 1 , Lung Diseases, Interstitial/diagnostic imaging , Lung/diagnostic imaging , Lung/metabolism , Neovascularization, Pathologic/diagnostic imaging , Fibrosis , Biomarkers , Biopsy , PrognosisABSTRACT
BACKGROUND: Although much progress has been made in diagnosis of carcinomas, no established methods have been confirmed to elucidate their morphological features. METHODS: Three-dimensional structure of esophageal carcinomas was assessed using transparency-enhancing technology. Endoscopically resected esophageal squamous cell carcinoma was fluorescently stained, optically cleared using a transparency-enhancing reagent called LUCID, and visualized using laser scanning microscopy. The resulting microscope images were converted to virtual HE images for observation using ImageJ software. RESULTS: Microscopic observation and image editing enabled three-dimensional image reconstruction and conversion to virtual HE images. The structure of abnormal blood vessels in esophageal carcinoma recognized by endoscopy could be observed in the 3 dimensions. Squamous cell carcinoma and normal squamous epithelium could be distinguished in the virtual HE images. CONCLUSIONS: The results suggested that transparency-enhancing technology and virtual HE images may be feasible for clinical application and represent a novel histopathological method for evaluating endoscopically resected specimens.