Zero Order


Publications and easy caffeine

Posted in Uncategorized by Amy Ross on February 19, 2012
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Hello there.  I haven’t had a whole lot to write about. I’ve been focusing on the job search and some home repair things that unfortunately, keep jumping in price. It’s so much fun to find out the previous owner installed something wrong!

I do have a little exciting news:  A paper I co-authored has been accepted for publication! The paper examines cell viability when encapsulated in superporous hydrogels. This paper was originally written by a previous Master’s student, but had been rejected for publication. I rewrote the intro and conclusion, and one of my labmates rewrote some of the methods and discussion in addition to improving the quality of the images.  “Macroporous Hydrogel Scaffolds: A Platform for Cell Encapsulation” will be published in Biomedical Materials in April.

In the news:

Pharmacy on a chip gets closer: Researchers at MIT performed a clinical trial with an implantable drug delivery device for osteoporosis patients. This is the first implantable drug delivery device I’ve read about that uses electronics; typically the release is governed by diffusion or degradation. Externally programmable pacemakers exist, so why not a drug delivery chip?

FDA to review inhalable caffeine: Aeroshot was actually developed by a bioengineering professor at Harvard. As a Diet Coke addict, I approve of this development 🙂  At $2.99 a tube, it’s cheaper than a cup of coffee at Starbucks.

 

Peptide release

Posted in Uncategorized by Amy Ross on August 15, 2010
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My latest experiments have involved measuring the release of MMP cleavable peptides from hydrogels. A short, simplified  breakdown:

The MMP cleavable peptide with a fluroescent molecule is conjugated to the hydrogel material monomer, PEGDA (polyethylene glycol diacrylate). The PEGDA is polymerized. Hydrogel samples are put in the presence of cells producing activated MMP-2.  The MMP should enter the hydrogel, cleave the peptide, and release part of the peptide (including the fluorescent molecule) into the cell media. By measuring the fluorescence in the cell media, the amount of peptide released can be determined. A control group is done where hydrogels are only in the presence of media (no cells), to verify that release is taking place.

I did the experiment above, measuring release at different time points up to 96 hours, but my results showed no significant difference in release between the cell group and the control group. My initial thought was that there were differences in conjugation.  Because I used a separate group of hydrogels for each time point, perhaps one group had more conjugated peptide than another. Tomorrow I’m going to try measuring release from the same group of hydrogels at different time points.

Another suggestion made by my PI is that the cells don’t produce active MMP when they become c0nfluent (covering the entire area of the growing surface). I’m using HT-1080s (a human fibrosarcoma cell line), which are known to produce active MMP-2. However, they divide very quickly. I did my experiment with a cell density of 500,00 cells/mL, and they became confluent within 24 hours of seeding.

I’m reviewing the thesis and lab notebooks of a previous student who did a lot of work on MMP-2 expression from cells.  I haven’t finished reviewing her experiments, but I noticed she used much lower cell densities than I am using.  A lower cell density is what I’ll attempt if the experiment I try tomorrow doesn’t show a significant release difference between experimental and control groups.

Other Uses for Hydrogels

Posted in Uncategorized by Amy Ross on July 12, 2010
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A few weeks ago, I talked a lot about using hydrogels for drug delivery. Today I’m going to talk about two other uses for hydrogels: one commercial, one experimental.

Use in Electrodes

Biological signals tend to be of low voltage, (e.g, 0.5-4 millivolts for cardiac potential), which I suppose is a good thing for us. The downside is that potentials measured from other sources-often referred to as noise-interfere with  the signal measurement. One of the problems with electrodes is that noise can occur if the electrodes slip out of place, or motion artifact.

Electrodes function by forming a series of opposite charges at the interface between the electrode and what it’s placed on. If the electrode moves, the charges on the other side of the interface are disrupted, and will have to redistribute to re-establish equilibrium.  During that redistribution, a potential difference forms, which will be measured and considered part of the signal.

Hydrogels that contain electrolytes (such as chlorine ions) are used as an interface between electrodes and the skin.  They keep the electrodes in place and reduce motion artifact.

Tissue Engineering

Tissue engineering is exciting stuff.  Can we grow skin, bone, or more complex body parts  from cells?  If we can, it would be a benefit to people with chronic wounds, burn patients, and (hopefully) people waiting for transplants. No doubt you’ve heard a lot about stem cells and the research done to get them to differentiate into a specific tissue type. One approach is to use a hydrogel as a scaffold, or support structure, for stem cells.

The properties of a hydrogel are similar to human tissues-they’re relatively soft and elastic. The premise is that the stem cells will take cues from the environment that will direct their differentiation. Additionally, hydrogels provide a 3-D environment more similar to the human body than the typical 2-D environment  in which cell studies are performed.

Our lab has approached this by using superporous hydrogels as a scaffolding material.  When the hydrogel is polymerized, the pores are created using sodium bicarbonate.  The pores that are created interconnect and allow stem cells to infiltrate the structure and proliferate.  One of the PhD students, Melanie, is currently working on this project. She’s using superporous hydrogels to differentiate human mesenchymal stem cells into bone tissue.

There’s way more than I could possibly say about the tissue engineering angle, but I’ll leave it at that for now.

Hydrogels

Posted in Uncategorized by Amy Ross on June 7, 2010
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The term hydrogel may be unfamiliar, but you’ve probably encountered its oldest and most successful commercial use: the soft contact lens. In order to understand hydrogels, first it’s necessary to explain a few things about polymers.

Polymers are material composed of long chains of repeating molecules. The molecule chains can be as long as tens of thousands of repeating units.  The properties of the polymer depend on the repeating unit and the circumstances under which it is made.  A variety of materials-plastic, rubber, and nylon to name a few-are made of polymers.

A hydrogel is a type of polymer that is  crosslinked and can absorb water. In crosslinking, the polymer chains  are bound together by another compound at regular intervals, creating a “mesh”. The molecular structure looks kind of like a fishing net:

Hydrogel structure at the molecular level

Hydrogels are useful in drug delivery because a drug placed in the hydrogel will diffuse out when placed in an aqueous environment. The rate of diffusion can be controlled by the mesh size (the size of the space bordered by the polymer chains and crosslinking compounds).

From "Matrix metalloprotease selective peptide substrates cleavage within hydrogel matrices for cancer chemotherapy activation". J. Tauro, B. Lee, S. Lateef, and R. A. Gemeinhart. Peptides, Volume 29, Issue 11 (2008): 1965-1973

The system I’m working with takes this idea one step further. It’s designed to treat glioblastoma multiforme, a particularly nasty form of brain cancer. The hydrogel is implanted at the tumor site at the time of resection. The drug in question is attached  to the polymer via a peptide. The peptide is the cleavage site for the enzyme MMP-2. MMPs are a class of enzymes that are produced in normal cells , but are overproduced in cancer cells. The MMP-2 enters the hydrogel and cleaves the peptide, thereby releasing the drug (see Figure).

My specific project is to demonstrate the MMP-2 actually penetrates the hydrogel, and doesn’t just cleave peptides on the surface. To accomplish this, I’ve made a two layer hydrogel. The bottom layer is polyethylene glycol diacrylate (PEGDA) and includes the MMP cleavable peptide.  The top layer is made of polyacrylamide.  Cells will be placed on the top layer. If the peptide is cleaved, we know that the MMP penetrated through the top layer and diffused through the bottom.

I’ve been working on this project for the past four months. I’ve created and optimized the hydrogel bilayer. In the past month, I’ve started cell experiments. I’m also working on some characterization of the hydrogels.

In the next few entries, I’ll be discussing some of the techniques I’ve been utilizing and challenges I have faced in the course of my research.