October 18, 2007
Howard Hughes Medical Institute
For sperm to penetrate an egg, they must first compress into a tight ball before springing into action. Researchers have now discovered a protein that can affect how DNA is packaged inside sperm so that they can scrunch up tightly enough to pierce the outer layer of the egg during fertilization.

The new studies in mice show that if this key protein is missing, DNA in sperm cannot be tightly packaged and the sperm will not be able to penetrate the egg. The researchers speculate that deficiencies in the protein may underlie some forms of male infertility.

“A small molecule that enhances the enzyme’s activity could be a useful fertility drug in cases where compromised function of the gene has caused infertility.”
Yi Zhang

The research team, which was led by Howard Hughes Medical Institute investigator Yi Zhang, published its findings online in Nature on October 18, 2007. Zhang and colleagues at the University of North Carolina at Chapel Hill collaborated on the studies with researchers in the Laboratory of Reproductive and Developmental Toxicology at the National Institutes of Health.

In their experiments, Zhang and his colleagues explored the function of the enzyme Jhdm2a, which is a histone demethylase. Histone demethylase enzymes activate genes by snipping molecules called methyl groups from histones. Histone proteins make up the “smart stuffing” in chromosomes—the core of proteins around which DNA winds so that it is packaged compactly.

Chemical modification of histones — such as the addition or subtraction of methyl groups – is an important mechanism for controlling the activation or repression of genes. This kind of epigenetic control mechanism is separate from other mechanisms that control gene expression, such as regulatory DNA elements that are embedded in the sequences of the genes themselves. Zhang’s research group is one of the leaders in establishing the role of demethylases in regulating gene activity.

The researchers focused on the function of Jhdm2a because their earlier studies had indicated that the gene for the protein is highly active in the testis. The protein also interested Zhang and his colleagues because Jhdm2a protein levels are highest during sperm maturation.

When the researchers knocked out the Jhdm2a gene in mice, they found that the animals’ sperm did not mature properly. On closer examination, they found that the genetic material, called chromatin, in the immature sperm of the knockout mice did not condense normally. Sperm chromatin must condense into a compact form in order for fertilization to be successful.

“In order for sperm to be able to enter the egg, the sperm chromatin has to be tightly packaged,” said Zhang. “It must become like a dense ball, so that when it hits the egg, it can penetrate. And in order for this DNA to be tightly packaged, the histone proteins must be replaced by other basic proteins.” The basic proteins include transition nuclear protein 1 (Tnp1) and protamine 1 (Prm1), said Zhang.

The researchers’ experiments established that the Jhdm2a demethylase specifically activates the Tnp1 and Prm1 genes. It does so by binding to the promoter region of the genes, which removes the methyl group that had been keeping the genes silent. Once the methyl group is removed, the Tnp1 and Prm1 genes are activated.

Zhang said that although their study was done in mice, it might well have implications for understanding some forms of human infertility. “It has been shown that there are many genes in mice that cause infertility when knocked out. But so far few of those genes has been found to be linked to human cases of infertility,” he said. “However, no one has paid much attention to these demethylase proteins. And since they play such a fundamental role in gene regulation in both mice and humans, there is a possibility that Jhdm2a plays a role in some types of human infertility.”

Zhang said that drugs that affect the Jhdm2a enzyme might have clinical use. “A small molecule that enhances the enzyme’s activity could be a useful fertility drug in cases where compromised function of the gene has caused infertility,” he said. “On the other hand, a small molecule that inhibits the enzyme’s activity could be a potential birth control drug.”

At Overlake Reproductive Health, passive 13.56 MHz RFID tags and interrogators track human sperm, eggs and embryos throughout the assisted-reproduction process.

By Claire Swedberg
RFID Journal
Copyright RFID Journal LLC 2008, Used With Permission

Oct. 15, 2007—Overlake Reproductive Health, located in Bellevue, Wash., has become the first reproductive-medicine center in the United States to deploy an RFID-based system for tracking human eggs, sperm and embryos. This system should help ensure that no identity mistakes are made during collection, storage and fertilization.

A female client can visit the clinic to be artificially inseminated by a partner’s sperm, or to have her egg fertilized in vitro (in a test tube) and then implanted in her uterus. For either procedure, the couple may worry that the sperm or egg might be accidentally switched with someone else’s, resulting in a baby that is not biologically theirs. Although such mistakes rarely happen, the experience can be traumatic for parents when they do occur, subjecting a clinic to lawsuits and negative publicity.

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Shaun Kelly

Until the RFID system was deployed in September, Overlake, like other in vitro fertilization (IVF) clinics, relied on the diligence of its employees to ensure that samples were never confused. When a patient provides sperm or eggs, the specimen is marked with the client’s name, and if transferred to another receptacle, it is again marked with that patients’ name. Usually, two employees manually check the names to prevent a mistake from being made. That system had been working appropriately, says Overlake’s laboratory director, Shaun Kelly, but patients were still uneasy.

At a recent American Society for Reproductive Medicine (ASRM) conference, Kelly happened upon Research Instruments, a U.K. manufacturer of RFID solutions, and saw a potential solution. “I was really intrigued by the whole thing from the get-go,” he says. Research Instruments provided Overlake with IVF Witness, an RFID-based system that helps keep specimens from being inadvertently switched.

Upon arrival, a patient is provided an ID card containing a 13.56 MHz RFID chip complying with ISO standard 15693. A staff member at the front desk inputs data about the patient, and a Research Instruments RFID interrogator captures the ID card number, which is linked to that data.

Each specimen is placed in a container with an RFID tag affixed to its bottom. When the container is placed on an interrogator, the system prompts the user to assign a particular patient to that specimen. IVF Witness permanently links the container’s tag ID number with that patient, so that the tag numbers for both sperm and eggs are input into the same patient account. When a specimen is sent to a lab, it passes several workstations, each equipped with an RFID reader. There are three readers in the sperm-prep lab; two in the embryology lab, where eggs are fertilized and developed; and one in the procedure room, where eggs are removed from a female client, and where sperm or embryos are implanted in the patient’s uterus.

At every step in the process, each specimen container is placed on a plate with an RFID reader, which captures the container’s tag ID. IVF Witness opens that patient’s account, and if any specimens tag IDs do not belong to that account, the system transmits an alert, emitting an audible alarm and displaying a red stop sign on the workstation screen. When this happens, the system cannot be restarted until an explanation is input to the system.

“The patients are incredibly happy with it,” Kelly says. New patients in particular, he adds—who have not yet had the opportunity to build a level of trust with the Overlake staff—find the RFID system reassuring. According to Kelly, Overlake continues with its original practice of hand-marking each specimen and using two witnesses to ensure the owner’s identity, but now it also has another layer of security. Although no situation has yet caused the system to issue an alert, Kelly says, the medical center has tested the system repeatedly and it is functioning properly.

“It is expensive,” Kelly says, citing the system’s price tag of nearly $60,000, and the clinic has had to extend some of that cost to clients. “They’re not complaining,” he notes. “They’re happy to have that security.”

Overlake typically completes up to 500 embryonic procedures annually. The system does not have FDA approval but doesn’t need it, according to Research Instruments. Still, the RFID system provider has tested it with mouse embryos to ensure that radio waves do not harm specimens. In Research Instruments’ tests, the tags transmitted continuously for four days without any perceptible effects on the mouse embryos.

RFID was chosen for this application, rather than bar-coding or some other technology, because it enabled a passive inventory check of the work area prior to the procedure being carried out, says David Lansdowne, technical director and patent holder at Research Instruments. With RFID, Lansdowne explains, lab personnel do not have to scan an item—they can simply place it on the workstation plate, and its ID number will be captured. Because automatic ID confirmation occurs as soon as a specimen is placed on the workstation, he says, a clinic “can ensure that laboratory SOPs [standard operating procedures] are being followed—something that is impossible with bar-code systems.”