Stomach Bugs Need Vitamin B6

H Pylori electron micrograph
Electron micrograph of H. pylori showing multiple flagella. Image: Wikimedia Commons - Yutaka Tsutsumi, M.D. Professor Department of Pathology Fujita Health University School of Medicine
Approximately half of the world’s population is infected with Helicobacter pylori, the bacterium that causes peptic ulcers and some forms of stomach cancer. Although ‘triple therapy’ with a proton pump inhibitor and two antibiotics – selected from a very limited number – can eradicate H.pylori, an increasing number of people are found to be infected with antibiotic-resistant bacteria. Scientists in Australia, New Zealand and France have now shown that H.pylori needs vitamin B6 to establish and maintain chronic infection, and have identified two genes in the vitamin B6 biosynthesis pathway as potential targets for new antibiotics.

The team used an established technique known as in vitro attenuation to create variants of a mouse-colonising strain of H.pylori with low infectivity and then compared the gene expression profiles of the attenuated bacteria with the original highly virulent strain. The most significant changes were found to be in the genes that encode homologues of the Escherichia coli vitamin B6 biosynthesis enzymes, PdxA and PdxJ, which catalyse sequential steps in the pathway. In vitro, H. pylori PdxA mutants could only be recovered when pyridoxal-5’-phosphate, the bioactive form of vitamin B6, was added to the growth medium whereas it was not possible to produce viable bacteria with mutated PdxJ. PdxA was also shown to be necessary for H. pylori to establish a chronic infection in mice.

Further studies showed that, in addition to its well known metabolic roles, vitamin B6 is needed for the synthesis of glycosylated flagella and for flagellum-based motility in H. pylori. The study, which is published in the new open access journal mBio, suggests that Pdx enzymes, which are present in a number of human pathogens, but not in mammalian cells, may present attractive targets for new antibiotic medicines.

Decoy Receptor Reverses Cancer Cachexia, Improves Survival in Mice

duck decoys
Image: Flickr - vhhammer
Debilitating muscle wasting or cachexia affects the majority of patients with advanced cancer but although the condition is believed to contribute to cancer-related deaths, the precise mechanisms by which cancer causes cachexia and those by which cachexia contributes to a poor prognosis are ill understood. There are currently limited treatment options for patients with cachexia, but scientists at Harvard Medical School and Amgen Research have now created a decoy receptor that can reverse cachexia in mice and increase survival, even though it has no effect on tumour growth.

ActRIIB is a high affinity activin type 2 receptor that mediates signalling by a subset of TGF-β family ligands, including myostatin, which inhibits muscle cell differentiation and growth, and activin, which is abundant in some cancer patients. Activation of ActRIIB initiates a signalling cascade that leads to increased degradation of myofibrillar proteins through the ubiquitin-proteasome pathway. In several mouse models of cachexia, administration of soluble ActRIIB (sActRIIB) was found not only to prevent further wasting but also to fully reverse both skeletal muscle loss and atrophy of the heart. Treatment with sActRIIB had no effect on fat mass or tumour growth and did not reduce elevated inflammatory cytokines, although it did stimulate feeding.

The study, which is published in the journal Cell, suggests that blocking the ActRIIB pathway has the potential to treat various muscle wasting diseases, particularly cancer cachexia, and if the results of the mouse studies translate to people, could also prolong the lives of cancer patients.

Never Mind the Colour – New Coating Kills MRSA on Contact

Paint pots
Image: Wikimedia Commons - Tpa2067
Methicillin-resistant Staphylococcus aureus (MRSA) infections are a particular problem in hospitals and other healthcare environments. MRSA can survive on normal surfaces and fabrics but researchers at Rensselaer Polytechnic Institute have now developed a coating that kills MRSA on contact. The coating contains lysostaphin linked by a short flexible polymer to carbon nanotubes and can be applied to surgical equipment, hospital walls, door handles and other surfaces. Lysostaphin is an example of a bacteriocin, a defensive bacterial antimicrobial agent that kills other, often closely related, bacteria. Lysostaphin, a cell wall-degrading enzyme, is produced by non-pathogenic strains of Staphylococcus bacteria and is very effective against Staphylococcus aureus, including MRSA, but completely harmless to humans and other organisms.

The lysostaphin-nanotube conjugate can be mixed with a wide range of surface finishes – in the present study, latex house paint was used. In tests, 100% of MRSA were killed within 20 minutes of contact with the paint. Treated surfaces can be washed repeatedly without losing their effectiveness and the team believe that the new coating is likely to prove superior to coatings that release biocides or those that ‘spear’ bacteria using amphipathic polycations and antimicrobial peptides. The team also believe that is unlikely that Staphylococcus aureus will be able to develop resistance to lysostaphin.

The study is published in the journal ACS Nano.

Nickel Allergy Linked to Innate Immune Response

piercing
Image: Wikimedia Commons - Nicor
Nickel allergy is one of the most common causes of allergic contact dermatitis and a team led by researchers at the University of Giessen have now shown that the response is linked to activation of a single receptor, toll-like receptor 4 (TLR4).

The family of toll-like receptors normally recognizes structurally similar molecules derived from microbial pathogens and plays a key role in host defense. The team showed that nickel directly activated human, but not mouse, TLR4 and studies with mutant proteins showed that non-conserved histidine residues at positions 456 and 458 were necessary for activation of human TLR4. Wild type mice do not show an allergy to nickel but transgenic mice expressing human, rather than mouse, TLR4 could be efficiently sensitized to the metal. This is the first time that an inorganic substance has been shown to activate this innate immune pathway and, since histidines 456 and 458 are not essential for responses to microbial lipopolysaccharide, the authors suggest that site-specific inhibition of TLR4 could provide a potential strategy for treatment of nickel allergy, which would not compromise normal immune responses. Nickel allergy affects millions of people and is often associated with earrings and jewellery for other body piercings.

The study is published in the journal Nature Immunology.

What’s the Structure of……??

chemicalized web page For those who haven’t encountered it yet, chemicalize.org is a tool for adding chemical information to your web browsing – and it’s free! Provided by the chemistry software company, ChemAxon, under a Creative Commons license, chemicalize can convert chemical names to structures – either on a query basis or converting an entire web page. In addition, ChemAxon have now added a page of calculated properties that can be accessed by clicking the generated 2D structure.

The tool will convert trivial chemical names such as saquinavir, as well as IUPAC names such as 7-chloro-1-methyl-5-phenyl-3H-1,4-benzodiazepin-2-one. Structure generation is based on ChemAxon’s name-to-structure software, although conversion of trivial names presumably relies on a database. I don’t know how comprehensive the database is, but it can certainly generate some interesting results when converting a web page – I hadn’t realised that trigger was a trivial name for something!

When a web page is converted, recognised structures are underlined in the text. Hovering over the underlined text produces a tooltip with the 2D structure of the molecule and this can be clicked to visit the calculated properties page. The properties page provides a variety of useful information including, logP, rotatable bond count, pKa etc. The layout of the properties page can also be adjusted by the user, with some standard layouts provided for medicinal or synthetic chemist.

You can see the chemicalized version of this post here and visit chemicalize.org for further information.

Converting Pancreatic α-Cells to β-Like-Cells

alpha beta There are four main cell types in the islets of Langerhans in the pancreas; α-cells which secrete glucagon, β-cells which secrete insulin, δ-cells which secrete somatostatin, and PP cells which secrete pancreatic polypeptide. Type I diabetes is an autoimmune disease in which the insulin-producing β-cells are destroyed and could potentially be treated by the creation of new β-cells, either from stem or stem-like cells or by conversion of another mature cell type. It has recently been shown that the transcription factor, Pax4, induces transdifferentiation of pancreatic α-cells into β-cells in adult mice and a team led by researchers at the Broad Institute of Harvard and MIT has now shown that a similar effect can be achieved with a small molecule.

BRD7389 structure
BRD7389
Using a mouse α-cell line, the team screened over 30,000 compounds and found that one of them, BRD7389, induced insulin expression after 3 days treatment. Induction of insulin gene expression in α-cells peaked at 5 days (ca 50-fold at a BRD7389 concentration of 0.85µM) and the cells adopted a β-cell-like morphology. Insulin protein levels were also increased from a basal α-cell state, although were much lower than the levels produced by mature β-cells. BRD7389 also increased insulin secretion in primary human islet cells although since there are many different cells types in the human tissue samples, it is also not possible to attribute the increase in insulin levels exclusively to conversion of α-cells to β-like-cells.

Follow-up studies suggested that upregulation of insulin expression potentially involved inhibition of multiple members of the RSK family of protein kinases, but more experiments are needed to fully elucidate the mechanism of action of BRD7389. The study demonstrates, however, that a small molecule can induce insulin expression in α-cells and suggests that such a strategy could be used to increase β-cell mass by transdifferentiation in vivo. The team now want to identify other small molecules that could be used to enhance the effects of BRD7389, and boost insulin production in people with type I diabetes.

The study is published in PNAS.

Blocking Tumour Suppressors May Allow Damage Repair

Salamander
Image: Flickr - Marshal Hedin
The ability to grow a replacement tail or limb, present in some species of amphibians such as salamanders and newts, has been lost in vertebrates. Earlier this year, scientists from the Wistar Institute and Washington University showed that mice lacking p21, a downstream target of the tumour suppressor p53, have a greater regenerative capacity than normal mice and now scientists at the Stanford University School of Medicine have shown the importance of other tumour suppression pathways in limiting regeneration in mammalian muscle cells.

Differentiated mammalian muscle cells are not normally able to divide but the team found that mouse myocytes can be induced to re-enter the cell cycle and begin proliferating by blocking the expression of two tumour suppressors, retinoblastoma protein (Rb) and ARF, a protein transcribed from an alternate reading frame of the INK4a locus. ARF is found in birds and mammals but not in lower vertebrates and, interestingly, is expressed at lower than normal levels in mammalian livers – the only organ with some regenerative capacity. When RNA interference was used to temporarily block expression of Rb and Arf in cultured mouse myocytes, the cells lost their differentiated properties, re-entered the cell cycle and began to proliferate. The cells are incorporated into existing muscle fibres when transplanted into mice, but only if Rb function was restored. Without functioning Rb, the new cells proliferated excessively and disrupted the structure of the muscle tissue.

Previous studies had shown that suppression of the Rb gene alone causes newt muscle cells, but not mammalian muscle cells, to re-enter the cell cycle.

Although knocking down tumour suppressor genes has obvious potential dangers, temporary silencing of gene expression may eventually allow regeneration of cardiac or pancreatic tissue if the technique is also successful in other cell types.

The study is published in the journal Cell Stem Cell.

Mouse T-cells Could Help in Fight Against Human Cancer

Mouse Biplane
Image: Wikimedia Commons - Wilfredor
T-cell receptors are integral membrane proteins that recognise foreign antigens and initiate a series of intracellular signalling cascades that allow the immune system to fight infection. To avoid autoimmune diseases, T-cells must be able to discriminate between ‘self’ and ‘foreign’ antigens but this discrimination may also prevent the immune system from recognising and destroying tumour cells.

Researchers led by a team from the Max-Delbrück-Center for Molecular Medicine have now developed transgenic mice that produce T-cell receptors that recognise human cancer cell antigens and could potentially be introduced into the T cells of cancer patients. Using embryonic stem cells loaded with human DNA, the team generated transgenic mice that express the entire human T-cell repertoire. Negative selection normally removes maturing T-cells that are capable of binding strongly to ‘self’ antigens but the mouse does not recognise human cancer cell antigens as ‘self’ and T-cells expressing receptors to these antigens are allowed to survive. T-cells with such high affinity receptors for cancer cell antigens are not produced in humans and the researchers hope that introducing the high affinity receptors into the T-cells of cancer sufferers will boost the immune system’s ability to recognise and destroy tumour cells. A first clinical trial to evaluate the efficacy and tolerability of the methodology in cancer patients is planned.

The study is published in the journal Nature Medicine.

Reprogramming a Heartbeat

heartbeat
Image: Flickr - Shannon Yeh
Once damaged, heart muscle has very limited capacity for regeneration but scientists at the Gladstone Institute of Cardiovascular Diseases have now discovered how to reprogram structural fibroblasts into functioning cardiac muscle cells (cardiomyocytes). The team explored the effects of transcription factors known to be important for development of the heart and found that a combination of just three (Gata4, Mef2c and Tbx5) was sufficient to rapidly and efficiently convert cardiac or dermal fibroblasts into contractile cardiomyocyte-like cells.

Gladstone scientists have previously converted mouse mesoderm – germ tissue from very early embryos – into cardiomyocytes and have reprogrammed adult cells into induced pluripotent cells which can then be converted into other cell types but, in the present study, adult cells have been directly reprogrammed into a different type of cell without involvement of a progenitor cell state. The team hope that going directly from one adult cell type to another might eliminate some of the perceived risks associated with the use of stem cells and that it will be possible to identify small molecules that are able to trigger the conversion. Although the technique has yet to be tested in human cells, and further refinement and characterisation of the reprogramming process will be needed, the heart has a large pool of fibroblasts which provide a potential source for regenerative treatments if they could be directly reprogrammed to beating cardiomyocytes.

The study is published in the journal Cell.

Low-Fat or Low-Carb?

diet
Image: Flickr – malias
There has long been debate about the relative merits of a low-carbohydrate diet, as popularised by Atkins, compared to the more traditional low-fat approach to weight loss. A low-carbohydrate diet has also been anecdotally associated with adverse effects on health.

A newly published clinical study, led by researchers at the Center for Obesity Research and Education at Temple University, Philadelphia, has now shown remarkably little difference between the two regimes. The study followed over 300 subjects randomly assigned to either diet over a two year period and, importantly, combined the diets with comprehensive behavioural treatment.

In the low-carb group, carbohydrate intake was limited to 20 g/d for 3 months in the form of low–glycemic index vegetables with unrestricted consumption of fat and protein. After 3 months, participants were allowed to increase their carbohydrate intake (5 g/d per wk) until a stable and desired weight was achieved. The low-fat diet consisted of limited energy intake (1200 to 1800 kcal/d) with less than 30% of the calories derived from fat. For the behavioural treatment, each participant attended group sessions weekly for the first 20 weeks of the study, every other week for the next 20 weeks, and once every other month for the remainder of the study. In each session, participants discussed topics such as goal setting, self-monitoring, and limiting triggers to overeating.

Although attrition was high at 2 years, there were no differences in weight, body composition, or bone mineral density between the groups at any time point. Weight loss was approximately 11 kg (11%) at 1 year and 7 kg (7%) at 2 years. The low-carbohydrate diet group had greater increases in high-density lipoprotein cholesterol (“good” cholesterol) levels at all time points, increasing by approximately 23% at 2 years, suggesting that a low-carb diet may have some cardiovascular benefit.

Gary Foster, Director of Temple’s Center for Obesity Research and Education and lead author of the study said:

When comparing these two popular weight loss plans, none of the existing research had included a comprehensive, long-term, behavioural support component. This research tells us that people wanting to manage their weight need to be less concerned with which diet they choose, and more concerned with incorporating behavioural changes into their plan.

The study is published in Annals of Internal Medicine.