Biomedical Engineering Reference
In-Depth Information
functionality of metabolic networks. By MFA, the intracellular metabolic fluxes
are calculated by the measurement of few extracellular metabolite concentrations
in combination with the stoichiometry of intracellular reactions. MFA is applied
to evaluate the intracellular metabolic conditions and to identify key metabolic
pathways or metabolites in the central metabolism. In MFA, mass balances over
all the intracellular metabolites are used in combination with the stoichiometry of
intracellular reactions to calculate the fluxes through the different branches of the
network. The intracellular fluxes are calculated by combining measurements of
extracellular metabolite concentrations, either with linear algebra or with linear
optimization. However, in any given scenario, completing the material balance is a
challenging task due to the possibility of multiple product formation.
7.3.4 IntracellularDegradation
Intracellular degradation of various materials in cells results from the action of a
combination of multiple mechanisms. Understanding intracellular degradation is
important to understanding the trafficking of metabolites during various disease
states and gene therapy. The use of various viral (such as adenoviruses and retro-
viruses) and nonviral (such as synthetic lipids and polymers) delivery systems to
deliver DNA for gene therapy applications (to correct genetic deficiencies or treat
acquired diseases) has shown that the efficiency of the transfection of nonviral
gene transfer is poor relative to viral vectors. Viral vectors have evolved mecha-
nisms to attach to cells, cross cellular membranes, evade intracellular transport
systems, and deliver their genomes into the appropriate subcellular compartment.
However, viral vectors have several restrictions such as limited DNA-carrying ca-
pacity, a lack of target-cell specificity, immunogenicity, and, most importantly, the
safety of the patient. Developing nonviral gene delivery carriers with a high trans-
fection efficiency is important for safe use, improving target-cell specificity, and
eliminating immunogenicity. Apart from concerns related to in the extracellular
inactivation and initial favorable interactions with the cell surface, intracellular
degradation is an important barrier. Hence, understanding the intracellular deg-
radation mechanism is very important for novel designs. There are three major
degradation pathways:
1. Lysosomes contain nearly 50 hydrolytic enzymes, including a variety of
proteases known as cathepsins. Lysosomes have an internal pH of near-
ly 5. If genetic material is introduced into a cell, a major concern is the
degradation of the foreign genetic material by the lysosomes. The inter-
nalization of the gene delivery system by endocytosis results in an endo-
somal traffi cking process, which, if not escaped, leads to degradation of the
therapeutic DNA in lysosomes. Hence, some strategies focus on evading
the lysosomal degradation. Nevertheless, the products of this activity are
monomers, which leave the lysosomes either via diffusion or with the aid
of specialized transport systems. The building blocks formed during deg-
radation are either reutilized for the biosynthesis of complex molecules or
further degraded to provide metabolic energy.
