Malaria and the Blood-Brain Barrier

 EPCR and ICAM Receptors

Neuromics Astrocytes and Pericytes were used in this important study-Yvonne Adams, Rebecca W. Olsen, Anja Bengtsson, Nanna Dalgaard, Mykola Zdioruk, Sanghamitra Satpathi, Prativa K. Behera, Praveen K. Sahu, Sean E. Lawler, Klaus Qvortrup, Samuel C. Wassmer, and Anja T.R. Jensen. (2021). Plasmodium Falciparum Erythrocyte Membrane Protein 1 Variants Induce Cell Swelling and Disrupt the Blood–Brain Barrier in Cerebral Malaria. Journal of Experimental Medicine, 218 (3). doi: 10.1084/jem.20201266.

This is the first study showing malaria pericytes interacting with human endothelial cells. Please note we have the complete BBB Model that can be used for studies like these.

Sex Differences on the Behavior of Human Brain Microvascular Endothelial Cells (HBMECS)

The Risk of Stroke
There are sex differences in risk for stroke and small vessel ischemic disease in the brain. Microvesicles (MV) derived from activated cells vary by cell of origin and the stimulus initiating their release. MV released from cells activated by inflammatory and thrombotic factors have the potential to disrupt endothelial cells of the brain microvasculature. Therefore, experiments were designed to identify sex differences in the phenotype of MV released from cultured human brain microvascular endothelial cells (HBMEC) in response to inflammatory and thrombotic stimuli (Biology of Sex Differences201910:26 https://doi.org/10.1186/s13293-019-0241-y).

Neuromics' HBMECS, derived from a male donor, were used to study the differences between Male and Female Cells. There are indeed striking differences in the expression of Cell Adhesion Molecules when the male and female cells were stimulated with inflammatory and thrombotic factors.

4 Fold increase in release of MV from male (open bars) and female (black bars) HBMEC expressing cell adhesion molecules following stimulation with either TNFα (20 ng/ml) or THR (2 U/ml) for 20 h

These molecules, in part, control tight junctions in HBMECS. In stroke patients, these are disrupted resulting in injury to the brain.

The findings  underscore the appropriateness of identifying the sex of cells used in research studies of MV disposition within the vasculature, as well as the sex of cells used to generate MV for in vitro studies

Human Primary Cells Trifecta

Tri-culture of Neuromics' Human Primary Cells in 3-D BBB Model

I mentioned in an earlier post that we take Researchers questioning the validity of our Human Primary Cells very seriously. We follow up with our clients to make sure our cells are working as expected. If they are not 100% happy, we offer a free replacement or full refund.

...So it is encouraging to see Researchers using our Human Pericytes,  Brain Microvascular Endothelial Cells (HBMES) and Astrocytes to build a static 3-D Blood-Brain Barrier Model.

Ece Bayir, M. Mert Celtikoglu, Aylin Sendemira. The use of bacterial cellulose as a basement membrane improves the plausibility of the static in vitro blood-brain barrier model. https://doi.org/10.1016/j.ijbiomac.2018.12.257.

Image: Pericytes, HBMECS, and Astrocytes 5 days after culturing. Live cells (green) and dead cells (red).

The investigators concluded, "Caffeine and sucrose permeability values obtained from all models were close to literature data and physiological values."

Human Brain Microvascular Endothelial Cell in Action

Recent Pubs
Our Human Brain Microvascular Endothelial Cells (HBMECs) are known for the performance in drug discovery and tox assays. These 21-CFR compliant cells are used in the "blood side" of our 3-D BBB Models.

1. Shavali Shaik, Bridget Kennis, Shinji Maegawa, Keri Schadler, Yang Yanwen, Keri Callegari, Rishi R. Lulla, Stewart Goldman, Javad Nazarian, Veena Rajaram, Jason Fangusaro, and Vidya Gopalakrishnan. (2018). REST upregulates gremlin to modulate diffuse intrinsic pontine glioma vasculature. Oncotarget. 2018 Jan 12; 9(4): 5233–5250. doi: 10.18632/oncotarget.23750 
2. Hu et al. (2016). Epigenetic Activation of WNT5A Drives Glioblastoma Stem Cell Differentiation and Invasive Growth. Cell. 167, 1281–1295. http://dx.doi.org/10.1016/j.cell.2016.10.039,
   

GREM-1 is required for tube formation in vitro (A) Q-RT-PCR analysis of GREM-1 gene expression in SU-DIPG-IV cells stably expressing either control shRNA or GREM-1-specific shRNA. Lentiviral constructs expressing two different GREM-1 shRNAs (GREM-1.1 and GREM-1.2) were used to knockdown GREM-1. Efficiency of GREM-1 knockdown was determined by Q-RT-PCR and expression was normalized to 18s RNA. Significance is as shown (**less than.01). (B-C) HUVEC or human brain microvascular endothelial cells (HBMEC) were cultured in endothelial cell medium and or conditioned medium from either control shRNA or shGREM-1.2 transfected SU-DIPG-IV cells. Tube formation in matrigel was measured after 16h and images were obtained. The ability of GREM-1 to rescue loss of tube formation upon REST knockdown was determined by addition of human-recombinant GREM-1 (rGREM-1) to conditioned media-endothelial media mix. Scale bars, 100μm. (C) Quantification of tubes in matrigel shown in Figure B (right panels). Data shown is mean +/- SD, ***p less than .001, n=3. (D) Western blot analysis to assess VEGFR2 levels in SU-DIPG-IV, -VI and –XIII cells, HUVECs and HBMECs was done using anti-VEGFR2 antibodies. Tubulin served as a loading control. (E) Western blot analysis was performed to assess AKT signaling downstream of GREM-1 interaction with its potential receptor VEGFR2 in HUVEC and HBMEC. Anti-pAKT (S473), anti-pAKT (T308), total AKT, and anti-actin were employed.

If these cells fit your assay requirements and need more data/info, do not hesitate to contact me, Pete Shuster, CEO at pshuster@neuromics.com or cell: 612-801-1007.

Neuromics' BBB Model and Permeability Assays

Crossing the Blood-Brain Barrier
Our BBB Model is being increasingly used by Bio-Pharma for drug permeability studies. Customers include Amgen, Genentech, Boehringer Ingelheim, and Merck.

These assays are key for making sure molecules/compounds of interest will cross the barrier into the brain and at what rate. This data, in part, add clarity on best candidates to move into in-vivo testing.

Human Blood Brain Barrier Model	3D45002	12 well
Human Blood Brain Barrier Model	3D45002	24 well
Human Blood Brain Barrier Model	3D45002	6 wells

I you have interest in using our models, Rose Ludescher, Manager of Customer Satisfaction, can provide information aligned with your assay requiremenst. rose@neuromics.com or 1-866-350-1500.

Neuromics Cornerstone

Human Primary + Stem Cells and 3-D Tissue Like Models

We are pleased by growing demand for our "hard to find" primary cells and 3-D BBB model. These are proving to be engines for drug discovery and toxicology assays.

We have have the capabilities isolate cells from healthy human donors. We then grow and characterize the cells and cryo-preserve in vials of 500,000 cells. They are now ready to culture.

We are increasingly determining types of cells to add to our growing catalog by meeting the specific demand of our clients. For example, our addition of primary human schwann cells and microglia happened, because they were needed for a specific client study.

Human Schwann Cells

Our 3-D Blood Brain Barrier Model is also working well in the hands of our clients. We are receiving positive input and this is supported by repeat orders. It is designed for users to test the ability of compounds, molecules and oligos to pass the BBB and penetrate into the brain.

Our cornerstone: hard to find human primary/stem cells + 3-D tissue like assays.

We can say, "Need cells or 3-D tissue like models?  Just ask us!". Pete Shuster, CEO and Owner-direct phone: 612-801-1007 or pshuster@neuromics.com

Need Cells From Diseased Donors?

Just Ask
Some of our customers have expressed the need for Cells that have the phenotype of a given disease. This is especially true for researchers doing drug discovery for neuro-diseases like Alzheimer's and Parkinson's.

Deriving "hard to find" cells from tissue is one of our cornerstones.

If you have need for any type of human cell(s), give us a shot. I can be reached at 612-801-1007 or pshuster@neuromics.com. Thank you. Pete Shuster,CEO and Owner, Neuromics.

"Hard to Find" Cells

You can Trust Your Results
Cells form the foundation of your cell based assays. We work hard to provide the cells you need and, more importantly that they are healthy and happy.

We are pleased to announce that we are recognized for having primary human cells that comply with federal and international regulations. This is especially important in drug discovery, because if the cells yield targets that result in future therapies this validation becomes increasingly valuable.

Human Brain Microvascular Endothelial Cells in Culture

Looking for "hard to find cells"? Check us out today.

Cancer-Associated Fibroblasts (CAFs) and Cells

Solutions for studying tumor growth, angiogenesis and metastasis.

Human Brain Astrocytes-New

Derived from Cortex

Immortalized Human Brain Microglia Cells

Isolated from healthy human brain tissue

Human Schwann Cells - New

Human spinal nerve derived primary cells

hN2™ Neuron Discovery Kits

Derived from H9 (WA09) ECSs-Consistent, Easy to Use & Cost Effective

Human alpha Motor Neurons Kit

hMSC Derived-includes 100 ml of media to optimize your cultures

Human Cardiomyocytes-New

Derived from hMSCs

Human Chondrocytes-New

Derived from hMSCs

Human Endothelial Cells-New

Artery, Microvascular, Vein & Umbilical Cord Derived + Culture Media

Human Fibroblasts-New

Pancreas and Neonatal Dermal Derived

NeuroNet Pure Human Neurons Discovery and HTS Kits

Pure Population of hPSC Derived Neurons

Human Osteoblasts-New

Derived from hMSCs

hMPro™ Human Mesenchymal Progenitors (hMPCs)

hESC Derived

Human Mesenchymal Stem Cells (hMSCs)

hMSCs Derived from Umbilical Cord Blood

Stressed Selected Stem Cells

They Survive to Thrive

hAstroPro Human Astrocyte+Astroglial-Neurons Co-Culturing Kits

Pure, Potent and Proven

Human Pericytes

Brain and Retinal Derived

3-D Cell Based Assay Solutions

Nanofibers, Hydrogels and Extracellular Matrix (ECM) Proteins

Need Human Cells? Ask Us!

From Astrocytes to Satellite CellsWe have many "hard to find" primary human cells in our catalog. We also have the ability to micro-dissect cells from both healthy and diseased tissue and make the related cells available in about 30 days. Here're are some of the currently available cells.

• Human Bladder Microvascular Endothelial Cells
• Human Cardiac Microvascular Endothelial Cells
• Human Dermal Lymphatic Microvascular Endothelial Cells
• Human Dermal Microvascular Endothelial Cells
• Human Glomerular Microvascular Endothelial Cells
• Human Large Intestine Microvascular Endothelial Cells
• Human Ovarian Microvascular Endothelial Cells
• Human Pancreatic Microvascular Endothelial Cells
• Human Prostate Microvascular Endothelial Cells
• Human Retinal Microvascular Endothelial Cells
• Human Synovial Microvascular Endothelial Cells
• Human Tonsil Microvascular Endothelial Cells
• Human Liver Sinusoidal Endothelial Cells
• Human Small Intestine Endothelial Cells

Human Breast CAFs

Images: aSMA Positive CAFs plus FC Data

We'll be posting more new cells here as they become available.

Pete Shuster-Owner and CEO--- pshuster@neuromics.com or 612-801-1007

3-D Cell Based Assays Are Evolving

3-D Multicell Models and Gels

There is a growing body of publications of why 3-D matters in drug discovery and toxicology assays. This table shows important distinctions for single cell assays.

Cellular characteristics	2D	3D	Refs.
Morphology	Sheet-like flat and stretched cells in monolayer	Natural shape in spheroid/aggregate structures	20,24,50
Proliferation	Often proliferate at a faster rate than in vivo	May proliferate at a faster/slower rate compared to 2D-cultured cells depending on cell type and/or type of 3D model system	17,51
Exposure to medium/drugs	Cells in monolayer are equally exposed to nutrients/growth factors/drugs that are distributed in growth medium	Nutrients and growth factors or drugs may not be able to fully penetrate the spheroid, reaching cells near the core	24,52
Stage of cell cycle	More cells are likely to be in the same stage of cell cycle due to being equally exposed to medium	Spheroids contain proliferating, quiescent, hypoxic and necrotic cells	18,24,53
Gene/protein expression	Often display differential gene and protein expression levels compared to in vivo models	Cells often exhibit gene/protein expression profiles more similar to those in vivo tissue origins	17,40,54
Drug sensitivity	Cells often succumb to treatment and drugs appear to be very effective	Cells are often more resistant to treatment compared to those in 2D culture system, often being better predictors of in vivo drug responses	17,33

Table: Key Differences in Cellular Characteristics and Processes in Two-Dimensional and Three-Dimensional Culture Systems

Assay Drug Dev Technol. 2014 May 1; 12(4): 207–218.

doi:  10.1089/adt.2014.573

From the table, we see, for the most part, the overall advantages of cells in 3-D for generating more in vivo like data. We also see potential issues like: "Nutrients and growth factors or drugs may not be able to fully penetrate the spheroid, reaching cells near the core".  

We have resolved penetration issues with our engineered Collagel Hydrogels. We have further enhanced the assays by including multicell assays that even more closely mimic the in vivo environment.

Our Blood Brain Barrier (BBB) Model, for example, includes our:

• Human Brain Microvascular Endothelial Cells
• Human Brain Microvascular Pericytes (HBMVPCs)
• Human Brain Astrocytes (HBA)

These cells are pre-cultured in the engineered gels and cost a fraction of the individual components (see: culturing and assay protocol).

Name	Catalog #			Size	Price
Human Blood Brain Barrier Model	3D45002-6			6 wells 12 well 24 well	$1,780 $2,600 $3,625

Our intention is for you to generate the most in vivo like data possible.

Solutions for Blood Brain Barrier Research

Neuromics Has the Key Components!

We are pleased to add Human Brain Microvascular Pericytes (HBMVPCs) to our solution set for Blood Brain Barrier (BBB) Researchers. These join our Human Brain Microvascular Endothelial Cells and hAstroPro Human Astrocyte+Astroglial-Neurons Co-Culturing Kits to round out our offerings of cells involved in BBB regulation.

 

Image: Human Brain Pericytes cultured with AlphaBioCoat (AC001).

Pericytes function at the BBB in at least two ways: by regulating BBB-specific gene expression patterns in endothelial cells, and by inducing polarization of astrocyte end-feet surrounding CNS blood vessels (see: http://www.nature.com/nature/journal/v468/n7323/full/nature09522.html).

Questions? Do not hesitate to call me or e-mail me-612-801-1007 or pshuster@neuromics.com). Pete Shuster CEO and Owner.