Shaking up cell biology

Elvis did it, Michael Jackson did it, and so do the mitochondria in our cells. They shake. While Elvis and Michael shook for decades before loud and appreciative audiences, mitochondrial oscillations have quietly bewildered scientists for more than 40 years.

Now, a team of scientists at US National Institutes of Health’s National Institute of Dental and Craniofacial Research (NIDCR) has imaged mitochondria for the first time oscillating in a live animal, in this case, the salivary glands of laboratory rats. The report, published online in the journal Cell Reports, shows the oscillations occur spontaneously and often in the rodent cells, which leads the researchers to believe the oscillations almost surely also occur in human cells.

“The movements could last from tens of seconds to minutes, which was far longer and frequently at a faster tempo than observed previously in cell culture,” said Roberto Weigert, Ph.D., an NIDCR scientist and senior author on the study. The mitochondria also appear to synchronize their movements not only in an individual cell but, quite unexpectedly, into a linked network of oscillators vibrating throughout the tissue.

“You look through the microscope, and it almost looks like a synchronized dance,” said Weigert. “The synchronization, to borrow an old cliché, tells us that we need to differentiate the forest from the trees – and vice versa – when studying mitochondria. It may be that the forest holds the key to understanding how mitochondria function in human health and disease.”

The mitochondrion is of one of several distinct compartments, or organelles, in the cell cytoplasm. Although mitochondria are jacks of many biochemical trades, they are best known as the power plants of the cell. They generate a continuous supply of the molecule ATP that, like bits of coal, serve as the cell’s main source of energy to power the heart to beat, muscles to stretch, and virtually every movement that the body makes.

To keep cells fully charged, mitochondria operate four biochemical production lines that coalesce with oxygen molecules from normal respiration to produce ATP. One of these production lines starts with processing the molecule nicotinamide adenine dinucleotide, or NADH. Weigert and colleagues recognized that they could use their high-magnification microscope to visualize NADH as it naturally emits electrons as part of the ATP production process.

The key was their choice of microscopy. Weigert and colleagues are masters of intravital microscopy, an extremely high-resolution technique that dates back to the 19th century. It had been too powerful to use in live animals until recently.

“Animals breathe, their hearts beat, and their appendages twitch,” said Weigert. “The combined effect under very high magnification is like watching a 6.0 earthquake. Everything shakes and blurs out of focus. We have developed approaches to better stabilize our organ of interest and minimize the motion artifacts. At this point, it is just a matter of generating more powerful optics to visualize the chemistry of life that really unfolds in the body, not under artificial laboratory conditions that stress cells and likely modify their behaviour.”

The powerful optics allowed the scientists to visualize the oscillations in their native milieu and to puzzle over their cause. Based on a series of subsequent experiments and observations, the researchers discovered that the oscillations are linked to the production of reactive oxygen species, a chemically interactive byproduct of making ATP. This finding suggests that the oscillations likely are not inherent to mitochondria but a response to conditions in their environment.

“These findings emphasize how important it is scientifically to study biology on its own terms, not under artificial laboratory conditions,” said Natalie Porat-Shliom, an NIDCR scientist and lead author on the paper. “We saw things in live animals that you don’t see in cell culture. The reasons, in this case, very well may be that the mitochondria continue to receive an influx of signals from the blood vessels, the nervous system, and their surrounding environment. The entire system can’t be reassembled in cell culture.”

Porat-Shliom noted that these findings should be of broad interest scientifically in framing studies of mitochondria, and may have future clinical implications. An estimated 2 million Americans have mitochondrial disease, an energy-depleting failure of mitochondria to function properly, which can have disabling effects on the brain, heart, kidneys, and other body systems. Many scientists also suspect that as mitochondria become better understood, they likely will be understood to play a more prominent role in human health and disease.

Link between breast implants and cancer under investigation

An international research group including Viennese pathologist Lukas Kenner has reviewed cases of possible association between breast implants and a form of lymphoma that may develop tumours at a later stage. The researchers conclude that breast implants can cause a new subtype of the rare yet malignant lymphoma known as ALCL. The research results have been published in the journal Mutation Research.

Worldwide there have been 71 documented cases of patients with anaplastic large cell lymphoma (ALCL) in which researchers suspected breast implants to be the cause. ALCL is normally found in the lymph nodes, as well as in skin, lung, liver and soft tissue, but not usually in the breast. Cases in which ALCL developed in the breast region almost exclusively involved patients who have had breast surgery. In these cases, ALCL developed around ten years after the operation. The tumours grew in the scar tissue around the implant.

Breast implants are generally safe and studies have found no association between breast surgery and other forms of cancer. ALCL itself is also an extremely rare occurrence. Among three million breast implants, there are between one and six reported cases of ALCL.

ALCL is divided into two subtypes. In one subtype, the cancerous cells produce an abnormal form of the protein ALK (anaplastic lymphoma kinase). The other type does not express ALK in tumour cells at all. While patients with ALK-positive lymphoma have a better chance of survival, the cancer is considerably more aggressive in ALK-negative cases.

Implant-related ALCL appears to form a third group. The cells do not express ALK, but patients have good survival rates. “This is a previously unrecognized, new subtype of ALCL,” Lukas Kenner explains. “We must now determine the exact causes behind its occurrence.”

The actual reasons why implants can cause lymphoma remain unclear. While some patients were successfully treated with chemotherapy and radiation therapy, the lymphoma in many cases subsided on its own following removal of the implant and the surrounding tissue. An abnormal immune response from the body could therefore be a cause of the cancer. Kenner and his team are now preparing for further studies in which implants and dentures will be examined in other parts of the body.

New discovery concerning glucose uptake in brown fat could help treat type 2 diabetes

Research findings that can likely be used to develop a new type of medicine for type 2 diabetes are published today in the Journal of Cell Biology. Researchers at Stockholm University have discovered a new mechanism that stimulates glucose uptake in brown fat – a tissue whose primary function is to generate heat by burning fat and sugar.

The major breakthrough of this discovery is of how glucose uptake is stimulated in brown fat. This knowledge can be used to pharmacologically stimulate this signalling pathway and lower blood sugar levels, which could lead to a cure for type 2 diabetes.

“One of the most interesting characteristics of this newly discovered signal pathway is that it differs from the signal pathway triggered by insulin. This means that the signal pathway in brown fat can most likely be activated even in patients with type 2 diabetes, where the insulin signalling is impaired,” says Professor Tore Bengtsson from the Department of Molecular Biosciences, the Wenner-Gren Institute, Stockholm University.

Although type 2 diabetes is a very serious and growing disease, there is still no definitive treatment or cure.

“Finding new ways to stimulate glucose uptake in tissues and thereby lower blood sugar levels is thus a matter of great interest,” says Prof Bengtsson.

Brown fat has been shown to be active in adults and is one of the tissues in the body that can be stimulated to the highest uptake of glucose per gram of tissue. Consequently, an increase in the uptake of glucose in brown fat can rapidly lower blood sugar levels.

“Our study shows that the body’s own stress hormones, epinephrine and norepinephrine, increase the uptake of glucose in brown fat. Epinephrine and norepinephrine can affect almost all our bodily organs by binding to receptors on the surface of a cell. We have shown how, and by what mechanism, adrenergic receptors found on brown fat stimulate the uptake of glucose. This is completely new and ground-breaking research,” says Prof Bengtsson.

Low carb diet shows benefits for diabetics compared to low fat diet

A low-carbohydrate diet has a good effect not only on blood glucose, but also on physical functions, bodily pain and general health, according to a diet study including patients with type 2 diabetes, which looked at the effects on blood glucose and blood lipids of a low-carbohydrate diet compared to a low-fat diet.

The results of a two-year clinical trial in patients with type 2 diabetes, led by Dr Hans Guldbrand, general practitioner, and Fredrik Nystrom, professor of internal medicine at Linköping University, was published in Diabetologia.

The 61 enrolled patients were randomly divided into two groups – one for each diet type and were expected to adhere to the respective diet throughout the study period. It was found that both diet-groups reduced weight equally but the effect on blood glucose was better in the low-carbohydrate group.

The effects of a low-carbohydrate diet and a low-fat diet on wellbeing have now been analysed in a study led by Associate Professor Margareta Bachrach-Lindström. A standardised analysis based on the SF- 36 questionnaire was performed. After 12 months in the trial, the low-carbohydrate group improved in regard to the physical component, which includes physical function, bodily pain and general health. No improvements were seen in the low-fat group, despite weight loss. Mental health was similar for both groups and remained unchanged during the study period and did not differ between the groups.

“The result is interesting; it provides an additional argument that a low-carbohydrate diet is beneficial in diabetes,” said Hans Guldbrand. “We also found no adverse effects on mental health with the low-carbohydrate diet, which an earlier study had indicated,” he added.

The interview with the study patients revealed that for both groups there were difficulties adhering to the diet when they ate elsewhere than at home. It could also be problematic if not all family members followed the same diet. Both groups expected health gains by adhering to given dietary advice. The low-carbohydrate diet group expressed that it could be difficult to refrain from potatoes and pasta. The diet for the low-fat group was described as relatively inexpensive and tasty. Benefits of the low-carbohydrate diet were that the patients felt less hungry and that their appetite for sweets disappeared.

doi: 10.1016/j.diabres.2014.08.032

Cause of donor death should not automatically exclude lungs from transplant consideration

Patients receiving lungs from donors whose cause of death was asphyxiation or drowning have similar outcomes and longterm survival as patients receiving lungs from traditional donors, according to a study in the October 2014 issue of The Annals of Thoracic Surgery.

Lungs from donors who died from asphyxiation or drowning are not routinely utilized because of potential damage sustained by the organs. However, the researchers show that lungs from donors whose cause of death was asphyxiation or drowning can be safely transplanted into patients with end-stage lung disease. They note that patient survival rates were not affected when lungs from cases involving asphyxiation and drowning were used.

They point out that if centres wanted to expand their individual criteria for donation, they could successfully expand their donor pool.

“For most patients with end-stage lung disease, transplant offers the only hope for survival, but there is a critical organ shortage, especially for patients on the lung transplant list. Increasing the potential donor pool would help reduce the number of patients who die while on the waiting list and help expand this lifesaving treatment to those who need it,” said Bryan A. Whitson, MD, PhD, from The Ohio State University Wexner Medical Center in Columbus. Dr Whitson and colleagues searched the United Network for Organ Sharing (UNOS) Standard Transplant Analysis and Research registry for lung transplants from 1987 to 2010 and assessed the association between donor cause of death and recipient survival, focusing on asphyxiation or drowning as the cause of death.

The researchers found 18,250 adult primary lung transplants, including 309 cases that involved asphyxiation or drowning. They also found that although the hospital stay was slightly longer (0.8 day) for recipients of lungs from asphyxiation or drowning deaths when compared with patients who received lungs from all other causes of donor death, survival rates were the same and there were no differences in treatment for rejection within the first year, post-transplant dialysis, or post-transplant stroke.

doi: 10.1016/j.athoracsur.2014.05.065

Study provides first evidence of longterm safety of transplanting human embryonic stem cells in humans

New research published in The Lancet provides the first evidence of the mediumterm to long-term safety and tolerability of transplanting human embryonic stem cells (hESCs) in humans.

hESC transplants used to treat severe vision loss in 18 patients with different forms of macular degeneration appeared safe up to 3 years post-transplant, and the technology restored some sight in more than half of the patients.

“Embryonic stem cells have the potential to become any cell type in the body, but transplantation has been complicated by problems including the risk of teratoma formation (a type of cancer that occurs when stem cells differentiate into multiple cell types and form incompatible tissues that can include teeth and hair) and immune rejection,” explained lead author Professor Robert Lanza, Chief Scientific officer at Advanced Cell Technology in the USA. “As a result, immunoprivileged sites such as the eye have become the first parts of the human body to benefit from this technology.”

In the two phase 1/2 studies, hESCs were differentiated into retinal pigment epithelium cells and transplanted into nine patients with Stargardt’s macular dystrophy and nine patients with dry atrophic age-related macular degeneration, the leading causes of juvenile and adult blindness in the developed world, respectively. No effective treatments exist for either condition, and eventually the photoreceptor cells of the retina degenerate leading to complete blindness.

All participants were injected with one of three different doses of retinal cells (50,000, 100,000 and 150,000 cells) into the subretinal space of the eye with the worse vision.

The hESC-derived cells were well tolerated for up to 37 months after transplantation. No safety concerns (eg. hyperproliferation or rejection) in the treated eyes were detected during a median follow-up of 22 months. Adverse events were associated with vitreoretinal surgery and immunosuppression, but none were deemed to be related to the hESC-derived cells.

Follow-up testing showed that 10 out of 18 treated eyes had substantial improvements in how well they could see, with 8 patients reading over 15 additional letters in the first year after transplant. Visual acuity remained the same or improved in seven patients, but decreased by more than 10 letters in one patient. Importantly, untreated eyes did not show similar visual improvements.

According to co-lead author Professor Steven Schwartz from the Jules Stein Eye Institute, Los Angeles, USA: “Our results suggest the safety and promise of hESCs to alter progressive vision loss in people with degenerative diseases and mark an exciting step towards using hESC-derived stem cells as a safe source of cells for the treat ment of various medical disorders requiring tissue repair or replacement.”

Writing in a linked Comment, Anthony Atala, Director of the Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, Winston- Salem, NC, USA says, “The work by Schwartz and colleagues is a major accomplishment, but the path to get to this point has not been smooth. Since the discovery of hESC in 1998, much has transpired, including political, ethical, and scientific debates, with an overall push to achieve the promise of human therapies. Now, we have follow-up that extends to longer than 3 years in patients treated with hESC-derived stem cells, showing both safety and apparent efficacy. Much work remains to be done before hESC and induced pluripotent stem cell therapies go beyond regulatory trials, but the path is now set in motion.”

doi: 10.1016/S0140-6736(14)61376-3

New test can help doctors choose best treatment for ovarian cancer

Researchers have devised a new test to help doctors diagnose ovarian tumours and choose the most appropriate treatment.

Successful treatment depends in part on accurately identifying the type of tumour, but this can be difficult. As a result, many women with cancer are not sent to the right specialist surgeon, or those with a benign cyst may have a more serious operation than they need.

In a study published in October 2014 the British Medical Journal, an international team led by Imperial College London and KU Leuven, Belgium describe a new test, called ADNEX, which can discriminate between benign and malignant tumours, and identify different types of malignant tumour, with a high level of accuracy.

The test is based on the patient’s clinical information, a simple tumour marker blood test and features that can be identified on an ultrasound scan. As well as identifying the type of tumour, the test expresses the confidence of the diagnosis as a percentage.

Doctors can use the test in a clinical database or by entering the patient’s details into a smartphone app, which was demonstrated to gynaecologists at the International Society for Ultrasound in Obstetrics and Gynecology World Congress in Barcelona in September 2014. The authors of the study say doctors could start using ADNEX straight away.

Professor Tom Bourne, from the Department of Surgery and Cancer at Imperial College London, said: “It’s very important to get the preoperative diagnosis right. If it isn’t right, the patient might have a more extensive operation than they need, for example having an ovary removed unnecessarily.

“If a tumour is benign, a woman might not need any treatment at all. If it is malignant, you need to know what type of tumour it is to choose the best treatment and that treatment needs to be carried out by specialist gynaecological cancer surgeon”

“At the moment, the way we assess women with ovarian cysts for the presence of cancer and select treatment lacks accuracy. This new approach to classifying ovarian tumours can help doctors make the right management decisions, which will improve the outcome for women with cancer. It will also reduce the likelihood of women with all types of cysts having excessive or unnecessary treatment that may impact on their fertility.”

The researchers developed the test using data from 3,506 patients from 10 European countries from 1999 to 2007, looking at which information available before the operation could be used to predict the diagnosis. They then tested the model on a further 2,403 patients between 2009 and 2012.

Studies have shown that ovarian cancer patients have a better chance of survival if they are referred to a specialised gynaecological cancer unit, but this only happens for a minority of women seeking treatment in the US and Europe.

Apart from the tumour type, the choice of treatment sometimes has to take into account implications for the woman’s fertility. Existing prediction models discriminate between benign and malignant tumours but lack accuracy and don’t sub-classify malignant tumours. The ADNEX model can discriminate between benign, borderline, stage I invasive, stage II-IV invasive, and secondary metastatic tumours.

The study was funded by the Flemish Government: Research Foundation - Flanders (FWO), Flanders’ Agency for Innovation by Science and Technology (IWT), iMinds and the National Institute for Health Research (NIHR) Imperial Biomedical Research Centre.

doi: 10.1136/bmj.g5920

New drug shows promise for treating spinal cord injuries

Injections of a new drug may partially relieve paralyzing spinal cord injuries, based on indications from a study in rats.

“We’re very excited at the possibility that millions of people could, one day, regain movements lost during spinal cord injuries,” said Jerry Silver, Ph.D., professor of neurosciences, Case Western Reserve University School of Medicine, Cleveland, and a senior investigator of the study published in Nature. Every year, tens of thousands of people are paralyzed by spinal cord injuries. The injuries crush and sever the long axons of spinal cord nerve cells, blocking communication between the brain and the body and resulting in paralysis below the injury.

On a hunch, Bradley Lang, Ph.D., the lead author of the study and a graduate student in Dr Silver’s lab, came up with the idea of designing a drug that would help axons regenerate without having to touch the healing spinal cord, as current treatments may require.

“Originally this was just a side project we brainstormed in the lab,” said Dr Lang. After spinal cord injury, axons try to cross the injury site and reconnect with other cells but are stymied by scarring that forms after the injury. Previous studies suggested their movements are blocked when the protein tyrosine phosphatase sigma (PTP sigma), an enzyme found in axons, interacts with chondroitin sulfate proteoglycans, a class of sugary proteins that fill the scars.

Dr Lang and his colleagues designed a drug called ISP to block the enzyme and facilitate the drug’s entry into the brain and spinal cord. Injections of the drug under the skin of paralyzed rats near the injury site partially restored axon growth and improved movements and bladder functions. “There are currently no drug therapies available that improve the very limited natural recovery from spinal cord injuries that patients experience,” said Lyn Jakeman, Ph.D., a program director at the US National Institutes of Health’s National Institute of Neurological Disorders and Stroke. “This is a great step towards identifying a novel agent for helping people recover.”

Initially, the goal of the study was to understand how interactions between PTP sigma and chondroitin sulfate proteoglycans prevent axon growth. Drugs were designed to mimic the shape of a critical part of PTP sigma, called the wedge. Different designs were tested on neurons grown in petri dishes alongside impenetrable barriers of proteoglycans. Treatment with ISP freed axon growth.

“It was amazing. The axons kept growing and growing,” said Dr Silver.

Next the researchers tested the potential of the drug on a rat model of spinal cord injury. For seven weeks they injected rats with the drug or a placebo near the site of injury. A few weeks later the rats that received the drug showed improvements in walking and urinating while the placebo treatments had no effect. The results suggested the drug passed into the brain and spinal cord.

When the researchers looked at the spinal cords under a microscope they found that the drug induced sprouting of axons that use the neurochemical serotonin to communicate. The sprouting axons were seen below the injury site. Treating some rats with a blocker of serotonin communication partially reversed the beneficial effects of ISP injections, suggesting the newly sprouting axons helped the rats recover.

The ISP drug did not cause spinal cord axons known to control movements to cross the scar and reconnect with brain neurons above the injury site. Dr Silver and his colleagues think this means the ISP-induced sprouting helped the rats recover by increasing the signal sent by the few remaining intact axons.

“This is very promising. We now have an agent that may work alone or in combination with other treatments to improve the lives of many,” said Dr Silver. He and his colleagues are seeking to test the ISP drug in preclinical trials. l doi: 10.1038/nature13974 ###

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