1906PeloNewsLiver/Islet

pelobiotech

Dive into your monthly PELONews and read some exciting scientific stuff as... ​ New Liver/Pancreas 2D/3D models for Cancer research, Drug Discovery & more Discover ECMs&Hydrogels for liver, pancreas &more #drugdiscovery #extracellularmatrix NEW Media for 2D/3D Human Hepatocytes Revolution of Enzyme Requirements for Human Islet Isolation Reducing the Cost of Human Islet Isolation DNs-Rh: fluorogenic substrate for GSTs Trabecular Meshwork Cells NEW Sales Contact in France

PELONEWS

WITH FOCUS ON LIVER&ISLETS

PELOBiotech Newsletter June/July 2019

“The heart has the nature of

knowledge, the liver of emotion,

the lungs of leaf..“

Dear Scientist,

the liver plays a critical role in human metabolism. As the gatekeeper of the digestive

track, this huge organ is responsible for drug breakdown and is therefor

the first to be injured due to overdose or misuse. Evaluating this drug-induced

liver injury is a critical part of pharmaceutical drug discovery and must be carried

out on human liver cells.

Pancreatic cancer is one of the most challenging research fields

today. Cancer develops when cells grow in an uncontrolled manner

and form masses or tumors in the pancreas. Due to the importance

of these organs, we have assembled this issue. Enjoy!

Christiane Büchsel

Editor in Chief PELONews

New Sales Contact in France

—Hildegard von Bingen,

German Nun & Healer-

Content

Featured Fields&Products




Liver/Pancreas 2D/3D models

Enzymes: Revolution of

Enzyme Requirements

for Human Islet Isolation

Reducing the Cost of Human

Islet Isolation

Especially for our French customers we have

an own dependance now for you in France.


NEW: 2D/3D Hepatocyte

Media

Pic. M.Vidal

I like to introduce to you M. Michel Vidal. He´s

representing our products and service to you and

very happy to talk to you about your research projects

and challenges.

M. Vidal is very experienced in the Life Science

sector and can support you in your daily business.

Please contact us or your personal contact in France

today.

M. Michel Vidal : Mobile + 33 (0)6 07 75 16 53



DNs-Rh: fluorogenic substrate

for GSTs

Trabecular Meshwork Cells

You like what you see? Sign

up for PELONews now at

www.pelobiotech.com or just

fill out the following form

online.


PELONews

Liver/Pancreas 2D/3D models

Go natural: Take native Hydrogels

Xylyx Bio has developed the native proteins and growth factors from the respective organ tissue for the cell culture and

thus reproduce a more natural environment. The ECM is available as coating, hydrogel or scaffold (custom order) for liver,

pancreas-and much, much more.

Pancreas: 2D Surface Coating , 3D Scaffold

Liver: 2D Surface Coating , 3D Hydrogel & Scaffold

Take your pick: Pancreas ECM cell culture substrates Exxtracellular matrix (ECM) is a major component of the cell microenvironment

with important signaling and regulatory functions. Derived from healthy acellular porcine porcine tissue, NativeCoat and

TissueSpec® pancreas matrix is rich in collagen types I, III, IV but also includes collagen type V.

TissueSpec® liver matrix is the organizational framework consisting of fibronectin and collagen types I, III, IV, V, and VI that mediates

hepatic endothelial and epithelial cell communication, modulates liver homeostasis, and supports repair and regeneration.

Excellent Growth Results for

Dog Primary Liver Organoids

The very innovative company Noviocell has developed Noviogel.

It is a synthetic matrix, free of animal and undefined components.

The hydrogel for cell culture behaves similar to collagen

or fibrin: Your Plus:

- functionalized with RGD peptides or Pure

- thermosensitive and reversible

But look for yourself! Already cultured successfully: Dog primary

liver organoids with Noviogel-RGD: Dog primary liver cells

were cultured in Matrigel and noviogel-RGD for 7 days. Liver

organoids cultured in noviogel-RGD showed excellent growth

characteristics, more organoids and bigger in size.

3D Co Culture Model for Liver, Pancreas Tumor Cells & more

Have a look at theese Applicable Cell Systems for Pancreas and Liver Tumor

cells. These cells have been tested sucessfuly for aggregate sphere formation

by our academic partners.

Full overview here: https://biopply.com/assets/Uploads/Applicable-Cell-

Systems.PNG

Details are described in the publication: A deep conical agarose microwell array for adhesion independent threedimensional

cell culture and dynamic volume measurement Andreas R. Thomsen et al., Lab Chip, DOI 10.1039/


Revolution of Enzyme Requirements for

Human Islet Isolation by Bob McCarthy

The quality of collagenase used for mammalian islet isolation has been a focus of clinical

research scientists soon after Ballinger and Lacy demonstrated that islet transplantation

could correct chemically induced diabetes in inbred strains of rats

(1). Clostridium histolyticum collagenase became a critical reagent because of the need

to isolate sufficient numbers of functional islets for islet transplantation in higher

mammals (e.g., dogs, pigs, humans). Typically, researchers evaluated several “lots” of

crude or enriched commercial collagenase products (i.e., traditional collagenase) for

their ability to recover pancreatic islets. The goal of lot qualification was to identify a

“good” lot of traditional collagenase that worked successfully in the islet isolation procedure

before purchasing larger amounts of a specific lot of product for routine use.

Good lots for human islet isolation were hard to find.

Camillo Ricordi’s development of a semi-automated method for human islet isolation

led him to focus on screening new collagenase lots once these lots were released for

sale by a supplier. During this process, Ricordi screened lots of Collagenase P, a crude

collagenase product first sold by Boehringer Mannheim Biochemicals in 1990. He

found that this material worked better for this application than competitive collagenase

products. One lot, lot 9, was used to isolate islets for many allo-islet transplantation

procedures (2). Collagenase P had higher collagenase and lower protease activity than

found in Sigma collagenase products used by many research scientists at that time.

Ricordi’s use of the Collagenase P product in clinical trials led to a meeting with R&D

management at Boehringer Mannheim Diagnostics in Indianapolis, Indiana about Boehringer

solving the “collagenase problem.” This discussion led to funding of a research

project at Boehringer Mannheim Biochemicals to identify and purify the key enzymes in

crude collagenase responsible for releasing islets from porcine or human pancreatic

tissue. The assumption was that the identification, purification, and formulation of purified

enzymes used at an appropriate dose would resolve the collagenase problem.

To read full paper, please visit: https://www.vitacyte.com/news/evolution-enzymerequirements-human-islet-isolation-link-recent-review/

Reference:

1 Ballinger WF, Lacy PE. Transplantation of intact pancreatic islets in rats. Surgery. 1972;72(2):175-86.

2 Ricordi C, Tzakis AG, Carroll PB, Zeng YJ, Rilo HL, Alejandro R, et al. Human islet isolation and allotransplantation

in 22 consecutive cases. Transplantation. 1992;53(2):407-14.

Enzymes

VitaCyte’s defined enriched

(DE) collagenase

products set a new

standard of consistency

and enables assessment

of a broader range of

collagenase: protease

ratios for cell isolation

than found with current

collagenase products.

DE Collagenase is

designed with increasing

amounts of enriched

collagenase added

to a fix amount of purified

BP protease.

Overview Rodent

Islet Overview discussion

of common

protocols used in rodent

islet isolations.

Includes a link to

a white paper that

summarizes a survey

of the affect of rodent

strain, age, sex, or

nutritional state on

expected islet yields

when purified enzymes

were used to isolate

islets. https://

www.vitacyte.com/

product-applications/

rodent-islet-applications


PELONews

Islet Isolation

Reducing the Cost of Human Islet Isolation

Clostridium histolyticum collagenase is a critical reagent for isolating islets from human pancreatic tissue. The development

of Liberaseä HI Purified Enzyme Blend overcame the problem of repetitive evaluation of different crude

collagenase lots to identify an acceptable product. The potential threat of transmission of spongiform encephalopathy

from bovine tissue led NIH to disallow the use of Liberase HI in the Clinical Islet Transplantation Consortium

(CITC) trial. Initially, Nordmark/Serva was chosen as the sole enzyme vendor for this trial, but VitaCyte and Roche

were added as suppliers after several experienced islet transplant laboratories were unable to achieve human islet

yields comparable to those obtained when using Liberase HI. All of the enzyme formulations in the CITC trial used

about 20 Wünsch Units of collagenase per g of trimmed tissue, the same dose used in the Immune Tolerance Network

Trial for human islet transplantation. Human islet isolation is an inefficient process since ≈ 53% of the islet isolations

had a sufficient number of islets for subsequent transplantation. The use of more consistent collagenase reagents

may increase the efficiency of this process.

A new era of understanding enzyme-mediated tissue dissociation began when Matsushita’s laboratory reported the

gene sequence and protein domain structure of C. histolyticumcollagenase. His laboratory showed class I (C1) and

class II (C2) collagenase were expressed by separate genes, validating earlier observations that showed these two

classes had different enzyme activity profiles and were separated by chromatographic techniques. They also showed

the degradation of native collagen only occurred with those molecular forms of collagenase that contained a catalytic

domain and at least one collagen binding domain.

VitaCyte correlated the specific collagen degradation activity (CDA) of purified natural collagenase to the molecular

form of the enzyme. Intact C1 with two collagen binding domains was 7-10% more active in degrading native collagen

when compared to “truncated” C1 or intact C2, each having only one collagen binding domain.

The new information on collagenase structure-function led to the development of a hypothetical model for collagenase-protease

digestion of tissue. The six key principles from this model are:

1. Differences in the efficiency of intact C1 vs truncated C1 or intact C2 likely reflects the ability of intact C1 with

two collagen binding domains to stick tighter to the collagen monomer during the degradation process than those

forms with a single collagen binding domain.

2. Collagenase appears to degrade collagen by moving from the amino terminal end of the collagen monomer to

carboxy terminal, degrading a collagen monomer in a fiber or fibril.

3. Excess collagenase CDA must be used to ensure rapid degradation of collagen; excess purified collagenase will

have a minimal adverse effect on cells since it has a narrow specificity for native collagen and gelatin.

4. Collagenase products that contain predominantly intact collagenase will have the highest specific CDA, and can

be used at lower doses in the islet isolation procedure than those products that contain multiple forms of

collagenase.

5. The ratio of C1 to C2 is unlikely to impact the efficiency of degradation of collagen in the extracellular matrix

since these two enzymes work cooperatively to degrade native collagen.

6. Choice and dose of neutral protease used for the isolation procedure is the critical variable that must be controlled

if excess intact collagenase is used in the isolation procedure.

Read full paper here: https://www.vitacyte.com/news/reducing-cost-human-islet-isolation


Human Islet

Read here common protocols used in human islet isolation.

Includes a table listing factors to consider when focusing

on enzymes used in these isolation procedures:

https://www.vitacyte.com/product-applications/humanislet-applications/

Hepatocytes

Rodent Hepatocytes Overview. Overview

discussion of methods for isolating rodent

hepatocytes. Includes a table of recommended

products and doses for use with rat

and mouse islet isolations.

Rodent HEPATOCYTE APPLICATIONS We

recommend: Defined Enriched Collagenases

DE Collagenase 60 or 600

DE Collagenase 10 or 100

These recommendations are dependent on

the protocol used in the isolation procedure.

(Recommended method: Seglen’s Two

-step Isolation Method 1,2.

F

To read more, please go to: https://

www.vitacyte.com/product-applications/

rodent-hepatocyte-applications/

1,2: Seglen P.O. (1976) Preparation of isolated rat liver

cells. Methods in Cell Biology 13, 29-83.;

2: Seglen PO. Isolation of hepatocytes. In: Celis JE, ed. Cell

biology : a laboratory handbook. San Diego :: Academic

For Hepatocytes:

Learn more


New Products

NEW: DNs-Rh:

DNs-Rh is a novel fluorogenic substrate for GSTs (Glutathione S-Transferases). Compared to conventional reagents,

DNs-Rh can be used for live cell imaging with high specificity and sensitivity discovered by Dr. Hiroshi Abe, Nagoya

University.

As many studies suggested, expression

level of GSTs is significantly

increased in cancer cells, GSTs are

considered as anti-cancer drugresistant

enzymes in malignant

cancer cells through the neutralization

of drugs. To understand biological

functions of GSTs, tools for

monitoring GST activity are very

important. Although several reagents

including CDNB (conventional

GSTs probe) for this purpose have

been developed, no tool to measure

intracellular GST activity was

commercially available.

DNs-Rh is a rhodamine 110 derivative which protected by DNs (dinitrobenzenesulfonamide).

This probe emits very low fluorescence (quantum yield = 0.0007) at normal state. After deprotected by thiols, rhodamine

110 is released and exhibits strong fluorescence (quantum yield = 0.645, S/N ratio ~900).

DNs-Rh can be used as a fluorogenic GST activity assay probe. An important advantage of this probe is high cellpermeability

and this probe is applied to intracellular GST activity under live cell condition.

As DNs group is a well characterized substrate for various types of GST members, DNs-Rh is able to monitor pan-

GST activity both in cell and in vitro.

More infos:

https://www.funakoshi.co.jp/exports_contents/81304

NEW:

These cells are endothelial-like cells in a sponge-like connective tissue located near the front

of the eye and are responsible for regualting eye pressure by controlling drainage of fluid into tubes that flow into

the bloodstream. Damage and dysfunction of TM have clinical significance. For instance, hypoxia increases DNA

methylation, accompanied by altered gene expression, whereas during the normal aging process, TM number decreases,

and senescent cells accumulate. In Glaucoma, a leading cause of irreversible blindness, decreased fluid

outflow causes an elevation of intraocular pressure and progressive loss of retinal ganglion cells. Physical changes

to the TM include increased fibrosis, fibronectin accumulation, and expression of ECM cross-linking enzymes.

Choose your Trabecular Meshwork Cells listed below or contact us for custom human or animal ocular cell types for

your research projects or applications: Ask Peter for more details sales@pelobiotech.com

Canine Trabecular Meshwork Cells: CnTMC, Human Trabecular Meshwork Cells: HnTMC, Porcine Trabecular

Meshwork Cells: PnTMC


More New Products

PELOBiotech‘s Premium-brand Cellovations®

launched NEW Media

Human Hepatocyte Medium - 2D (HHM-2D) and Human Hepatocyte

Medium - 2D (HHM-3D)

HHM-2D and HHM-3D are complete media formulated for optimal cultivation of all hepatocytes,

human and animal, etiher to grow in 2D after plating or to build spheroids (3D).

They are serum-free and growth factor containing culture media and contain essential and

non-essential amino acids, vitamins, organic and inorganic compounds, hormones, trace

minerals together with the growth factor containing supplements. They contain NO FCS. The

media are delivered as Kit with separate supplements. The media may be used for functionel

long-term cultivation of hepatocytes to study hepatotocxity of chemicals and drug-drug

interactions, or in hepatocyte bioreactors. Please be aware that hepatocytes do not proliferate.

They may be used to study hepatotocxity of chemicals and drug-drug interactions, or in hepatocyte

bioreactors. Please ask Lothar for further Details.

Cat. #: PB-MH-053-5990-2D or PB-MH-053-5990-3D


PELONews

PELONews: Refer a friend

Save

this date

We are convinced

that we convince

you with our expertise

and

excellent service.

So if you are already

a PELO-

Fan, please

recommend us to

a colleague and

friend – you will

get a nice thankyou

personalized

reward. More details?

Call us now.

Ipita: Meet us at the

IPITA 2019 July, 02-

05 July, 2019 in Lyon,

France. You may

meet our CEO Dr.

Peter Frost at the

booth of our partner

Vitacyte at the

7 th World Congress of

the

INTERNATIONAL

PANCREAS & ISLET

TRANSPLANTation.

Association. It´s a big

family meeting, so join

us now and meet Vitacytes‘s

CEO Bob

McCarthy at booth#8.

Testimonials: All that matters is you!

We are so happy, that you are our customer. And even more if we

may use your words to be recommended to other customers and

scientists: Because we can say a lot about us —but all that matters

is you.

Enjoy your daily coffee or pot of tea with this great and unique

cup—made with love and only for you. Send us your favorite color

(and if you like a favorite short citation)and you will have this gift

asap on your desk. Thank you.

How to

reach us

If you need any

further assistance or if

you like what you see,

tell us:

PELOBiotech GmbH

Klopferspitz 19

82152 Planegg |

Germany

Tel.: +49 89 517 286

59 0 |

info@pelobiotech.com

www.pelobiotech.com

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comply to the new GDPR guidelines.

Managing Directors:

Dr. Peter Frost,

Dr. Lothar Steeb

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