Sweet hopes for diabetics Good news for diabetics.

Sweet hopes for diabetics Good news for diabetics.

A three-year study carried out by Universiti Teknologi Malaysia in Skudai has confirmed previous findings that cinnamon has the potential to lower sugar levels.

UTM research and development manager Prof Dr Mohammad Roji Sarmidi said their research showed that the spice, known as kayu manis locally, has positive effects on the disease, especially Type II diabetes.
Type II diabetes causes cells to lose their ability to respond to insulin, the hormone that tells the body to remove excess glucose from the bloodstream. This condition usually develops in people in their middle age and prematurely kills an estimated 100 million of the world’s population every year. Dr Mohammad Roji said herbalists all over the world had used cinnamon in the treatment of diarrhoea and arthritis, as cinnamon extract was found to improve blood circulation, heal wounds, reduce pain spasm and prevent ulcer and allergies.
“In the last decade, laboratory studies have also revealed that cinnamon extract mimicked insulin action in the cells,” he said. Insulin regulates glucose metabolism, helping body cells to convert glucose to energy and keep blood sugar levels normal. “Studies by the Agriculture Research Service in the United States have also found that certain substances in cinnamon helps cells become more responsive to insulin,” he added. Cinnamon is an ingredient used in cooking, and in cakes, pastries and beverages like coffee and tea.
Dr Mohammad Roji said UTM would conduct further studies next year, which would cover tests on animals and metabolic profiling for diabetic patients.

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Hybridoma - a technique for eternal production of monoclonal antibodies in cell cultures

An explanation of the diversity of the immune system
The most important task for the immune system is to defend the body against bacteria, virus and other microorganisms. The specific defense is exerted by a subgroup of white blood cells (lymphocytes). The immune system needs to recognize and react specifically with a large number of foreign substances (antigens). How the lymphocytes develop these vital properties and how they build up the highly specialized recognition system of the immune apparatus has long been an area of intensive research. pharmaguideline.
Niels K. Jerne is the leading theoretician in immunology during the last 30 years. In three main theories he has elucidated central issues concerning specificity, development and regulation of the immune system in a comprehensive and convincing way. By his theories Jerne has outlined the development of modern immunology.
Theory 1: Specificity is predeterminedIn his Natural-Selection
Theory of Antibody Formation from 1955 Jerne explains the development of a specific antibody response in the following way. Each individual has a large number of natural antibodies with specificities for all antigens towards which the individual can respond. These antibodies develop already during fetal life in the absence of external antigens. The foreign antigen then selects the antibody molecule which has the best fit. The antigen-antibody binding stimulates the production of this particular antibody specificity.Jerne's natural-selection theory contrasted to the dogmatic views of the antibody response as formulated in the instruction theories which were prevailing at that time. According to these theories the antigen serves a template for the production of antibodies.In Jerne's natural selection theory it is implied that the generation of the enormous number of antibody specificities is independent of exogenous antigens. This view on the nature of the immune system constitutes the basis for modern immunology.
--->Regulatory affairs in pharmaceutical <---- Theory 2: Reactivity against self-antigens creates diversity
The natural-selection theory is mainly concerned with the maturation of the immune system after it has acquired the ability to react with antigen. In the second theory on the Somatic Generation of Immune Recognition set forth in 1971 Jerne explains how the immune system develops from stem cells to mature lymphocytes which can react with antigen. He presupposes that every individual possesses all genes needed for the production of antibodies, and antibody-like molecules, which can bind all strong transplantation antigens of the species. Jerne suggests that lymphocytes mature in the thymus gland and in other lymphoid organs where they are exposed to the transplantation antigens of the individual. Cells which recognize the antigens are stimulated and enter cell division. As mutations accumulate in rapidly dividing cells new immunological specificities may develop. At the same time the specificities of the lymphocytes for self transplantation antigens are weakened. The mature lymphocytes will recognize foreign antigen associated with transplantation antigens. The theory explains how the immune system normally matures through the influence of self antigens. It also offers an explanation for the regulation of immunological specificity by genes belonging to the transplantation system.
Theory 3. Antibodies, anti-anti-bodies ...In his third main theory, the Network Theory from 1974, Jerne explains how the specific immune response is regulated. The theory has greatly stimulated research and led to new insights into the immune system. Recently its principles have been applied to diagnosis and treatment of disease.
A basis for the network theory was the observation that antibodies can elicit anti-antibodies directed against antigen binding structures on the first antibody .Moreover, anti-antibodies can stimulate the production of still another generation of antibodies, anti-anti-antibodies. Essentially, this antibody cascade is endless successively adding new specific properties to the immune system. The various antibody generations either stimulate or suppress the production of one another. Under normal conditions the network is balanced. When an antigen is introduced the equilibrium is disturbed. The immune system tries to restore balance which leads to an immune response against the antigen.1. Infectious diseases. Anti-antibodies have been used in animals as a kind of vaccine against parasitic infections (trypanosomiasis), urinary tract infections, hepatitis and other infectious diseases.
2. Allergy. Anti-pollen antibodies may elicit allergic symptoms when an allergic person is exposed to pollen. The production of anti-pollen antibodies has been prevented in animals by anti-antibodies.
3. Autoimmune disease. Autoimmune disease may be caused by antibodies directed against the body's own tissues. Experimental autoimmune disease has been successfully treated with anti-antibodies.
4. Transplantation. Anti-antiimmunity may be important in organ transplantation by contributing to immunological tolerance against antigen on the foreign graft.
5. Endocrinology. Anti-antibodies against hormones and hormone receptors may prevent binding of the hormone to the receptors. This has been described for insulin and its receptor.
6. Tumours. Anti-antibodies have been attempted as treatment of certain tumours of the human immune system.
--->Pharmaceutical Regulatory affairs <---- Hybridoma - a technique for eternal production of monoclonal antibodies in cell cultures
Besides gene technology, which has already been honoured by several Nobel Prizes, the hybridoma technique represents the most important methodological advance within the field of biomedicine during the 1970s. The development of this technique is based on several observations concerning basic biological phenomena.
There are cells in the body - immune lymphocytes - which can produce millions of different antibodies. However, each single cell can only produce antibodies with a certain predetermined specificity. A prerequisite for the formation of a multitude of antibodies is, therefore, the existence of an excess of lymphocytes. If the body is exposed to a certain foreign antigen there may be stimulation of a lymphocyte which fortuitously has been endowed with the capacity to identify this particular antigen. This lymphocyte then starts to divide and forms a clone of cells which produces identical - monoclonal - antibodies.
The development of a clone of cells in connection with a normal immune response occurs under carefully controlled conditions. In rare cases, however, the body loses control over a clone of antibody producing cells. This may lead to formation of a special type of tumour (myeloma). Myeloma cells usually retain their capacity to produce a certain antibody, but because of the accidental emergence of the tumour one normally does not know with which antigen this antibody reacts.
White blood cells responsible for producing antibodies are highly specialized cells. As a consequence they lack capacity to survive for a longer time if they are removed from the body and incubated in a tissue culture medium. In contrast, myeloma cells can occasionally be cultivated continuously. Since long, biomedical research workers have nourished the dream to be able to propagate clones of cells which produce antibodies with predetermined specificity. This dream materialized when Georges J.F. Köhler and César Milstein in 1975 introduced the so-called hybridoma technology for production of monclonal antibodies. The principle features of the hybridoma technology is as follows pharma .blogspot.com
Figure 2. Principle steps in the production of a hybridoma. Spleen cells are prepared from animals, usually mice, which have been immunized with a selected antigen. These cells are then fused with myeloma cells maintained in culture in the laboratory. The product of this fusion is referred to as a hybridoma. Surprisingly, a hybrid of two cells can survive and also continue to divide. In this particular hybrid the myeloma cells contribute the capacity for survival, whereas the spleen cells direct the synthesis of antibodies with the preselected specificity. By special arrangements it is possible to achieve a multiplication of hybridoma cells but not of isolated myeloma cells. The hybrids obtained are propagated in a highly diluted state so that colonies deriving from single hybrid cells can be isolated. By use of a sensitive method the clones which produce the specific antibodies are identified. A particular hybridoma can then be used for future, unlimited production of a highly specific antibody.
The availability of monoclonal antibodies has opened completely new possibilities for basic as well as applied biomedical research. The following examples of the use of monoclonal antibodies can be given.
1. Detailed studies of the distribution of different functions in different parts of antigen molecules. These studies may concern building elements of infectious agents; cell products such as enzymes and hormones; surface structures of cells etc. The mapping of variations in the surface components of influenza virus which explain the occurrence of repeated infections is one example.
2. High degree purification of substances, e.g. interferon, by taking advantage of the unique capacity displayed by a particular monoclonal antibody to bind to a certain antigen. In this case one uses a technique referred to as affinity chromatography.
3. Diagnostic characterization of diseases by identification of special structures on the surface or on the inside of cells. Hereby it is possible to distinguish between different forms of tumours and follow the development of tumours. Furthermore, it is possible to distinguish between different kinds of normal white blood cells. This is of importance for the characterization of certain immune deficiency conditions as seen e.g. in connection with the disease AIDS (acquired immune deficiency syndrome).Diseases caused by infectious agents can also be diagnosed by use of monoclonal antibodies. Thus, virus infected cells and bacteria or parasites inside or outside cells can be identified with a unique degree of specificity.
4. Treatment of AIDS diseases. Monoclonal antibodies against specialized white blood cells have been used with some success in connection with transplantation. There may also be possibilities to use monoclonal antibodies for treatment of tumours.

Recombinant DNA Technology in the Synthesis of Human Insulin

Recombinant DNA Technology in the Synthesis of Human Insulin
Since Banting and Best discovered the hormone, insulin in 1921. diabetic patients, whose elevated sugar levels are due to impaired insulin production, have been treated with insulin derived from the pancreas glands of abattoir animals. The hormone, produced and secreted by the beta cells of the pancreas' islets of Langerhans, regulates the use and storage of food, particularly carbohydrates.
Although bovine and porcine insulin are similar to human insulin, their composition is slightly different.Consequently, a number of patients' immune systems produce antibodies against it, thus neutralising its actions and resulting in inflammatory responses at injection sites. Added to these adverse effects of bovine and porcine insulin, were fears of long term complications ensuing from the regular injection of a foreign substance, as well as a projected decline in the production of animal derived insulin.These factors led researchers to consider synthesising Humulin by inserting the insulin gene into a suitable vector, the E. coli bacterial cell, to produce an insulin that is chemically identical to its naturally produced counterpart.
This has been achieved using Recombinant DNA technology.
This method is a more reliable and sustainable method than extracting and purifying the abattoir by-product.Understanding the genetics involved.
The structure of insulin.
Chemically, insulin is a small, simple protein. It consists of 51 amino acid, 30 of which constitute one polypeptide chain, and 21 of which comprise a second chain. The two chains are linked by a disulfide bond.
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Inside the Double Helix.
The genetic code for insulin is found in the DNA at the top of the short arm of the eleventh chromosome. It contains 153 nitrogen bases (63 in the A chain and 90 in the B chain).DNA Deoxyribolnucleic Acid), which makes up the chromosome, consists of two long intertwined helices, constructed from a chain of nucleotides, each composed of a sugar deoxyribose, a phosphate and nitrogen base. There are four different nitrogen bases, adenine, thymine, cytosine and guanine.The synthesis of a particular protein such as insulin is determined by the sequence in which these bases
are repeated .
Insulin synthesis from the genetic code.
The double strand of the eleventh chromosome of DNA divides in two, exposing unpaired nitrogen bases which are specific to insulin production .
The role of the mRNA strand, on which the nitrogen base thymine is replaced by uracil, is to carry genetic information, such as that pertaining to insulin,from the nucleus into the cytoplasm, where it attaches to a ribosome
The nitrogen bases on the mRNA are grouped into threes, known as codons. Transfer RNA (tRNA) molecules, three unpaired nitrogen bases bound to a specific amino acid, collectively known as an anti-codon pair with complementary bases (the codons) on the mRNA. The Vector (Gram negative E. coli).
A weakened strain of the common bacterium, Escherrichia coli (E. coli), an inhabitant of the human digestive tract, is the 'factory' used in the genetic engineering of insulin. When the bacterium reproduces, the insulin gene is replicated along with the plasmid, a circular section of DNA . E. coli produces enzymes that rapidly degrade foreign proteins such as insulin. By using mutant strains that lack these enzymes, the problem is avoidedIn E. coli, B-galactosidase is the enzyme that controls the transcription of the genes. To make the bacteria produce insulin, the insulin gene needs to be tied to this enzyme. Inside the genetic engineer's toolbox
Restriction enzymes, naturally produced by bacteria, act like biological scalpels, only recognising particular stretches of nucleotides, such as the one that codes for insulin.
This makes it possible to sever certain nitrogen base pairs and remove the section of insulin coding DNA from one organism's chromosome so that it can manufacture insulin . DNA ligase is an enzyme which serves as a genetic glue, welding the sticky ends of exposed nucleotides together.
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Manufacturing Human insulin.

The first step is to chemically synthesise the DNA chains that carry the specific nucleotide sequences characterising the A and B polypeptide chains of insulin The required DNA sequence can be determined because the amino acid compositions of both chains have been charted. Sixty three nucleotides are required for synthesising the A chain and ninety for the B chain, plus a codon at the end of each chain,signalling the termination of protein synthesis. An anti-codon, incorporating the amino acid, methionine, is then placed at the beginning of each chain which allows the removal of the insulin protein from the bacterial cell's amino acids.
The synthetic A and B chain 'genes' are then separately inserted into the gene for a bacterial enzyme, B-galactosidase, which is carried in the vector's plasmid. At this stage, it is crucial to ensure that the codons of the synthetic gene are compatible with those of the B-galactosidase. The recombinant plasmids are then introduced into E. coli cells. Practical use of Recombinant DNA technology in the synthesis of human insulin requires millions of copies of the bacteria whose plasmid
has been combined with the insulin gene in order to yield insulin. The insulin gene is expressed as it replicates with the B-galactosidase in the cell undergoing mitosisThe protein which is formed, consists partly of B-galactosidase, joined to either the A or B chain of insulin . The A and B chains are then extracted from the B-galactosidase fragment and purified. Biological implications of genetically engineered Recombinant human insulin. Human insulin is the only animal protein to have been made in bacteria in such a way that its structure is absolutely identical to that of the natural molecule. This reduces the possibility of complications resulting from antibody production. In chemical and pharmacological studies, commercially available Recombinant DNA human insulin has proven indistinguishable from pancreatic human insulin. Initially the major difficulty encountered was the contamination of the final product by the host cells, increasing the risk of contamination in the fermentation broth. This danger was eradicated by the introduction of purification processes. When the final insulin product is subjected to a battery of tests, including the finest radio-immuno assay techniques, no impurities can be detected. The entire procedure is now performed using yeast cells as a growth medium, as they secrete an almost complete human insulin molecule with perfect three dimensional structure.
This minimises the need for complex and costly purification procedures. The issue of hypoglycaemic complications in the administration of human insulin.
Since porcine insulin was phased out, and the majority of insulin dependent patients are now treated with genetically engineered recombinant human insulin, doctors and patients have become concerned about the increase in the number of hypoglycaemic episodes experienced. Although hypoglycaemia can be expected occasionally with any type of insulin, some people with diabetes claim that they are less cognisant of attacks of hypoglycaemia since switching from animal derived insulin to Recombinant DNA human insulin.(16) In a British study,
published in the 'Lancet", hypoglycaemia was induced in patients using either pork or human insulin, The researchers found "no significant difference in the frequency of signs of hypoglycaemia between users of the two different types of insulin."
An anecdotal report from a British patient who had been insulin dependent for thirty years, stated that she began experiencing recurring, unheralded hypoglycaemia only after substituting Recombinant DNA human insulin for animal derived insulin. After switching back to pork insulin to ease her mind, she hadn't experienced any unannounced hypoglycaemia. Eli Lilly and Co., a manufacturer of human insulin, noted that a third of people with diabetes, who have been insulin dependent for over ten years, "lose their hypoglycaemic warning signals, regardless of the type of insulin they are taking." Dr Simon P. Wolff of the University College of London said in an issue of Nature , "As far as I can make out, there's no fault (with the human insulin)." He concluded, "I do think we need to have a study to examine the possible risk.

Sterile filtration

Sterile filtration

Clear liquids that would be damaged by heat, irradiation or chemical sterilization can be sterilized by mechanical filtration. This method is commonly used for sensitive pharmaceuticals and protein solutions in biological research. A filter with pore size 0.2 µm will effectively remove bacteria. If viruses must also be removed, a much smaller pore size around 20 nm is needed. Solutions filter slowly through membranes with smaller pore diameters. Prions are not removed by filtration. The filtration equipment and the filters themselves may be purchased as presterilized disposable units in sealed packaging, or must be sterilized by the user, generally by autoclaving at a temperature that does not damage the fragile filter membranes. To ensure sterility, the filtration system must be tested to ensure that the membranes have not been punctured prior to or during use.
To ensure the best results, pharmaceutical sterile filtration is performed in a room with highly filtered air (HEPA filter) or in a laminar flow cabinet or "flowbox", a device which produces a laminar stream of HEPA filtered air.

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