“Research is better when both sexes are included”

Ivana Jaric, what exactly is “sex as a biological variable” – and why does it matter?

Ivana Jaric: The biological sex can have a significant impact on how the body functions, how diseases develop and how treatments work. But historically, many studies, especially in biomedical research, used only male animals or didn’t analyze the data by sex. That means a lot of potentially important differences between males and females were overlooked. About ten years ago, the U.S. National Institutes of Health introduced the Sex as a Biological Variable (SABV) Initiative to encourage researchers to consider biological sex as a key variable in their studies.

Does this scientific oversight also apply to in vitro research, such as cell culture or organoids?

Jaric: Yes, very often. The sex of cells used in cell culture experiments has traditionally been ignored, despite evidence that it can influence critical cellular behaviors. For insance, only 25% of studies published in leading cardiovascular journals even reported the sex of the cells – and most of them used male cells.

Sex refers to biological differences (such as chromosomes, hormones and reproductive organs) while gender involves social and cultural identities, roles and expectations. In animal research, particularly with rodents such as mice and rats, experiments focus on biological sex.

Would you argue that SABV impacts scientific rigor and reproducibility?

Jaric: Let’s say you’re developing a new antidepressant and only test it in male animals. It might look promising in preclinical testing, but then fail in human trials – where women are often overrepresented because they have higher rates of depression. That’s because sex affects brain structure and function, drug absorption, metabolism and appropriate dosing. Ignoring these differences increases the risk that the treatment won’t work as expected in half the population, which can lead to failed trials, wasted funding and delays in getting effective treatments to patients.

So, yes, the research is better when both sexes are included. We can detect sex-specific effects that might otherwise be missed. It leads to more accurate conclusions and better reproducibility. It makes science more relevant, more efficient and more reflective of real-world populations.

And now you’re creating a pan-European SABV network – and you’ve even received an EU Cost Action Grant to support this endeavor.

Jaric: I want to help build a strong, connected, interdisciplinary community in Europe, one that supports researchers, shares best practices and ultimately improves the quality and impact of research when both sexes are included.

My motivation for submitting the EU-SABV COST Action came from a very simple reason: when researchers are now asked to include animals of both sexes in preclinical studies, they often don’t know how. There’s a lack of clear tools and guidelines – especially across different scientific disciplines.

Have you observed differences in the use of male and female lab animals across scientific disciplines?

Jaric: For example, in neuroscience, the field I work in, there used to be a strong male bias: for every six studies conducted on male animals, only one included females. In pharmacology and physiology, male outnumber females almost four to one. But in immunology and cancer research, it’s often the opposite: female animals are preferred because they’re less aggressive and easier to group-house.

However, all scientific fields can benefit from the inclusion of both sexes. Doing so helps us understand the biological mechanisms that may increase or decrease risk in one sex compared to the other.

Another issue is that many researchers confuse the concept of SABV with conducting studies specifically designed to investigate sex differences. But SABV simply means including both sexes – not necessarily focusing on the differences.

But doesn’t including both sexes double the number of animals used?

Jaric: That’s a common concern, but it’s a misconception. Including both sexes doesn’t mean you have to double your sample size. Instead of using eight males per group, you can use four males and four females. You still have the same number of animals overall. But, if we want to detect sex differences after an intervention or treatment, the total number of animals usually needs to be increased by about 30%. But this is worthwhile, because it gives us the opportunity to develop more effective, tailored medical interventions

This approach improves generalizability and actually aligns better with the 3R principles (replacement, reduction, refinement). We forget that using only one sex can lead to findings that aren’t broadly applicable. And since both sexes are naturally born in labs, it makes scientific and ethical sense to use both.

The European Cooperation in Science and Technology (COST) is a funding organization for the creation of research networks, called COST Actions. These networks offer an open space for collaboration among scientists across Europe (and beyond) and thereby give impetus to research advancements and innovation. COST Actions are typically made up of researchers from academia, SMEs, public institutions and other relevant organizations or interested parties.

How will you launch the SABV network in Europe?

Jaric: We’ll start by listening. A comprehensive survey of researchers across Europe will help us to understand what they feel is missing. Based on those results, we’ll design educational activities – webinars, summer schools, symposia and workshops – with both theoretical and practical elements.

For example, the theoretical workshops might focus on statistics and study design, while the practical ones could cover how to monitor hormone levels in male and female animals, how to model menopause or andropause in rodents, how to determine sex in vitro – or even how to properly handle and house both sexes in the lab. These are the kinds of details that often get overlooked but are essential for good research.

Starting in October this year, we have four years to develop evidence-based guidelines and offer training that actually helps researchers.

You draw on expertise from disciplines like statistics, bioinformatics and veterinary science. Why is this interdisciplinary approach so important?

Jaric: To really get the most out of this initiative, we need strong communication and collaboration across disciplines. It’s not just helpful, it’s absolutely essential. No single field can address all the challenges on its own.

For example, working with statisticians is critical, especially when designing studies that include both male and female animals. Without that support, we risk having underpowered studies that may not produce meaningful or generalizable results.

Veterinarians and animal welfare experts are also key partners. Applying the 3R principle means designing studies that are both ethically sound and scientifically robust. Sometimes, using animals of both sexes in a single, well-structured experiment is more informative than repeating studies with just one sex, which may produce results that don’t translate broadly.

What role does bioinformatics play?

Jaric: Collaboration with bioinformaticians is crucial, but comes with analytical challenges. Some genes are located on the X or Y chromosomes, and others are influenced by sex hormones like estrogen, testosterone and progesterone. These hormones affect not just reproduction but also behavior and immune responses. So, we need bioinformatics experts to help us interpret this complex data and establish clear guidelines for how to account for sex in these kinds of studies.

How will you connect SABV research with clinical practice?

Jaric: We’re reaching out to clinicians to make sure updates to preclinical guidelines also apply in clinical contexts. The EU-SABV COST Action is mainly focused on basic and preclinical research, but there are important clinical efforts – like the Chair for Gender Medicine at the University of Zurich. By connecting these efforts at a European level, we can take a unified approach and build a stronger bridge between the lab and the clinic.