Estimation of absolute number of alveolar epithelial type 2 cells in mouse lungs: a comparison between stereology and flow cytometry

Objective Iatrogenic laryngotracheal stenosis (iLTS) is the pathologic narrowing of the glottis, subglottis, and/or trachea secondary to intubation or tracheostomy related injury. Patients with type 2 diabetes mellitus (T2DM) are more likely to develop iLTS. To date, the metabolomics and phenotypic expression of cell markers in fibroblasts derived from patients with T2DM and iLTS are largely unknown. Study Design Controlled in vitro cohort study. Setting Tertiary referral center (2017-2020). Methods This in vitro study assessed samples from 6 patients with iLTS who underwent surgery at a single institution. Fibroblasts were isolated from biopsy specimens of laryngotracheal scar and normal-appearing trachea and compared with controls obtained from the trachea of rapid autopsy specimens. Patients with iLTS were subcategorized into those with and without T2DM. Metabolic substrates were identified by mass spectrometry, and cell protein expression was measured by flow cytometry. Results T2DM iLTS-scar fibroblasts had a metabolically distinct profile and clustered tightly on a Pearson correlation heat map as compared with non-T2DM iLTS-scar fibroblasts. Levels of itaconate were elevated in T2DM iLTS-scar fibroblasts. Flow cytometry demonstrated that T2DM iLTS-scar fibroblasts were associated with higher CD90 expression (Thy-1; mean, 95%) when compared with non-T2DM iLTS-scar (mean, 83.6%; P = .0109) or normal tracheal fibroblasts (mean, 81.1%; P = .0042). Conclusions Scar-derived fibroblasts from patients with T2DM and iLTS have a metabolically distinct profile. These fibroblasts are characterized by an increase in itaconate, a metabolite related to immune-induced scar remodeling, and can be identified by elevated expression of CD90 (Thy-1) in vitro.

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Immature platelet values indicate impaired megakaryopoietic activity in neonatal early-onset thrombocytopenia

Thrombosis and Haemostasis ◽

10.1160/th09-03-0148 ◽

2010 ◽

Vol 103 (05) ◽

pp. 1016-1021 ◽

Cited By ~ 27

Author(s):

Hannes Hammer ◽

Christoph Bührer ◽

Christof Dame ◽

Malte Cremer ◽

Andreas Weimann

Keyword(s):

Bone Marrow ◽

Flow Cytometry ◽

Intensive Care ◽

Early Onset ◽

Absolute Number ◽

Severe Thrombocytopenia ◽

Platelet Counts ◽

Immature Platelet Fraction ◽

Neonatal Thrombocytopenia ◽

Immature Platelets

SummaryNewly released platelets, referred to as immature platelets, can be reliably quantified based on their RNA content by flow cytometry in an automated blood analyser. The absolute number of immature platelets (IPF#) and the immature platelet fraction (IPF%) reflect megakaryopoietic activity. We aimed to analyse the implication of these parameters in analysing the pathomechanism of early-onset neonatal thrombocytopenia. Platelet counts and IPF were determined at day 1 to 3 (d1 to d3) in 857 neonates admitted to intensive care. In thrombocytopenic patients (platelet counts<150 x 109/l, n=129), IPF# was significantly lower (8.5 ± 2.7 x 109/l), than in non-thrombocytopenic neonates (9.5 ± 3.6 x 109/l, n=682, p<0.05). IPF% was significantly higher in thrombocytopenic (9.3 ± 7.9%) vs. non-thrombocytopenic neonates (4.1 ± 1.8%, p<0.001). While neonates with early-onset infection (n=134) had lower platelet counts (199 ± 75 x 109/l) compared to controls (230 ± 68 x 109/l, n=574, p<0.01), there were no differences in IPF# or IPF%. Likewise, “small for gestational age” infants (SGA, n=149) had lower platelet counts at d1 (199 ± 75 x 109/l, p<0.001) than controls, but no differences in IPF. A trend towards lower IPF# was detected if SGA infants with platelet counts <100 x 109/l (5.4 ± 3.9 x 109/l, n=11) and thrombocytopenic neonates with infection (9.9 ± 7.3 x 109/l, n=10, p=0.11) were compared. The evaluation of IPF# indicates that thrombocytopenia in neonates is likely due to a combination of increased platelet consumption and inadequate megakaryopoietic response by the neonatal bone marrow. Furthermore, SGA neonates with moderate and severe thrombocytopenia might have a pronounced suppression of megakaryopoiesis compared to neonates with infection.

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Absolute number of circulating CD34+ cells is abnormally low in refractory anemias and extremely high in RAEB and RAEB-t; novel pathologic features of myelodysplastic syndromes identified by highly sensitive flow cytometry

Leukemia Research ◽

10.1016/s0145-2126(99)00167-8 ◽

2000 ◽

Vol 24 (2) ◽

pp. 163-174 ◽

Cited By ~ 17

Author(s):

Kengo Fuchigami ◽

Hiroyuki Mori ◽

Tatsuki Matsuo ◽

Masako Iwanaga ◽

Kazuhiro Nagai ◽

...

Keyword(s):

Flow Cytometry ◽

Myelodysplastic Syndromes ◽

Absolute Number ◽

Pathologic Features ◽

Cd34 Cells ◽

Highly Sensitive

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Measuring Biofouling Potential in SWRO Plants with a Flow-Cytometry-Based Bacterial Growth Potential Method

Membranes ◽

10.3390/membranes11020076 ◽

2021 ◽

Vol 11 (2) ◽

pp. 76

Author(s):

Nirajan Dhakal ◽

Sergio G. Salinas-Rodriguez ◽

Joshua Ampah ◽

Jan C. Schippers ◽

Maria D. Kennedy

Keyword(s):

Flow Cytometry ◽

Bacterial Growth ◽

Potential Method ◽

Absolute Number ◽

Growth Potential ◽

Added Value ◽

Feed Water ◽

Dissolved Air Flotation ◽

Ultra Filtration ◽

Pre Treatment

Measuring the bacterial growth potential of seawater reverse osmosis (SWRO) feed water is an issue that is receiving growing attention. This study developed and demonstrated the applicability of the flow-cytometry (FCM)-based bacterial growth potential (BGP) method to assess the biofouling potential in SWRO systems using natural microbial consortium. This method is relatively fast (2–3 days) compared to conventional bioassays. The effect of the potential introduction of nutrients during measurement has been studied thoroughly to achieve the lowest measure value of about 45,000 cells/mL, which is equivalent to about (10 µg-C glucose/L). The BGP method was applied in two full-scale SWRO plants that included (i) dissolved air flotation (DAF) and ultra-filtration (UF); (ii) dual-media filtration (DMF) and cartridge filter (CF), which were compared with the cleaning frequency of the plants. A significant reduction (54%) in BGP was observed through DAF–UF as pre-treatment (with 0.5 mg Fe3+/L), while there was a 40% reduction by DMF–CF (with 0.8 mg Fe3+/L). In terms of the absolute number, the SWRO feed water after DAF–UF supports 1.5 × 106 cells/mL, which is 1.25 times higher than after DMF–CF. This corresponds to the higher cleaning-in-place (CIP) frequency of SWRO with DAF–UF compared to DMF–CF as pre-treatment, indicating that the BGP method has an added value in monitoring the biofouling potential in SWRO systems.

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Differences in Type 1 and Type 2 intracytoplasmic cytokines, detected by flow cytometry, according to immunosuppression (cyclosporine A vs. tacrolimus) in stable renal allograft recipients

Clinical Transplantation ◽

10.1034/j.1399-0012.1999.130506.x ◽

1999 ◽

Vol 13 (5) ◽

pp. 400-409 ◽

Cited By ~ 40

Author(s):

Lionel Rostaing ◽

Olivier Puyoo ◽

Jean Tkaczuk ◽

Christine Peres ◽

Anne Rouzaud ◽

...

Keyword(s):

Flow Cytometry ◽

Cyclosporine A ◽

Renal Allograft

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Increased LHX9 expression in alveolar epithelial type 2 cells of patients with chronic obstructive pulmonary disease

Respiratory Investigation ◽

10.1016/j.resinv.2021.08.007 ◽

2021 ◽

Author(s):

Koji Okutomo ◽

Naoya Fujino ◽

Mitsuhiro Yamada ◽

Takuya Saito ◽

Yoshinao Ono ◽

...

Keyword(s):

Chronic Obstructive Pulmonary Disease ◽

Pulmonary Disease ◽

Chronic Obstructive ◽

Obstructive Pulmonary Disease ◽

Alveolar Epithelial

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GATA6 regulates differentiation of distal lung epithelium

Development ◽

10.1242/dev.129.9.2233 ◽

2002 ◽

Vol 129 (9) ◽

pp. 2233-2246 ◽

Cited By ~ 1

Author(s):

Honghua Yang ◽

Min Min Lu ◽

Lili Zhang ◽

Jeffrey A. Whitsett ◽

Edward E. Morrisey

Keyword(s):

Gene Expression ◽

Epithelial Cell ◽

Surfactant Protein ◽

Gata Factor ◽

Lung Epithelium ◽

Alveolar Epithelial ◽

Distal Lung ◽

Transgenic Embryos

GATA6 is a member of the GATA family of zinc-finger transcriptional regulators and is the only known GATA factor expressed in the distal epithelium of the lung during development. To define the role that GATA6 plays during lung epithelial cell development, we expressed a GATA6-Engrailed dominant-negative fusion protein in the distal lung epithelium of transgenic mice. Transgenic embryos lacked detectable alveolar epithelial type 1 cells in the distal airway epithelium. These embryos also exhibited increased Foxp2 gene expression, suggesting a disruption in late alveolar epithelial differentiation. Alveolar epithelial type 2 cells, which are progenitors of alveolar epithelial type 1 cells, were correctly specified as shown by normal thyroid transcription factor 1 and surfactant protein A gene expression. However, attenuated endogenous surfactant protein C expression indicated that alveolar epithelial type 2 cell differentiation was perturbed in transgenic embryos. The number of proximal airway tubules is also reduced in these embryos, suggesting a role for GATA6 in regulating distal-proximal airway development. Finally, a functional role for GATA factor function in alveolar epithelial type 1 cell gene regulation is supported by the ability of GATA6 to trans-activate the mouse aquaporin-5 promoter. Together, these data implicate GATA6 as an important regulator of distal epithelial cell differentiation and proximal airway development in the mouse.

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