When an academic team in 2023 claimed to have found lymphatic vessels, tiny tubes in the body that carry immune cells and drain fluid from tissues, inside mouse and human bones, other biologists were intrigued. The discovery contradicted a widely held belief that bone was one of the few places these vessels didn’t penetrate. The group’s study, published in Cell, also suggested the tubes help bone regenerate after injury—a finding with obvious medical relevance.
Now, a team of scientists report today in Cell that they see no evidence of lymphatic vessels in healthy bone, and poke holes in the previously published experiments. The authors of the original 2023 paper reject the criticisms in a simultaneously published rebuttal. The warring papers have had a rocky path to publication: The journal made an eleventh-hour decision to halt their posting online last month because of concerns about allegations in the rebuttal aimed at some of the critique’s authors. The rebuttal has since been heavily re-edited.
Some researchers not involved in any of the three papers say they’re pleased to finally see concerns about the 2023 study publicly aired by Cell, and are inclined to believe the criticisms. The critics make a “strong case,” says Graeme Birdsey, a vascular scientist at Imperial College London. “The fact you’ve got all these groups coming together to disprove the [2023] paper … and [that] people have not been able to reproduce it convincingly, suggests that there’s definitely issues there.”
Lymphatic tubes have long been documented on the outside edges of healthy bone. But before 2023 they’d only been conclusively found deep inside bones in people with rare conditions such as Gorham-Stout disease, in which skeletal invasion by lymphatic vessels leads to dramatic bone loss.
That appeared to change when University of Oxford cell biologist Anjali Kusumbe and colleagues noticed cells bearing protein markers associated with lymphatic endothelial cells (LECs), which line lymphatic vessels, deep inside bones of normal mice. 3D images of these and human bones revealed LECs arranged in tubelike structures, and gene expression analyses of human bone marrow provided further evidence for the LECs’ presence, the researchers reported. In living mice, the team also showed that killing LECs with toxins impaired the animals’ ability to repair radiation-injured bones. The 2023 work was “paradigm-shifting,” Kusumbe tells Science, and could lead to ways to re-generate bones in people with fractures.
Still, some in the scientific community, including Kusumbe’s former postdoc adviser, Ralf Adams at the Max Planck Institute for Molecular Biomedicine, were skeptical the study captured true lymphatic vessels. To see for themselves, he and his colleagues sought to replicate the findings, later joining forces with cell biologist Bin Zhou at the Chinese Academy of Sciences, who already had a similar effort underway.
They and other collaborators soon concluded that the 3D imaging techniques and gene expression analyses used in the 2023 paper didn’t easily distinguish between lymphatic vessels outside and inside the bone. And the markers used to identify LECs could tag other cells, too, their data suggest. Adams says the original experiments destroying LECs in living mice weren’t persuasive either, in part because the toxins likely killed additional cell types, making the animals generally unwell and perhaps better explaining their impaired bone healing.
The critics’ own experiments looking at normal bone, which added different genetic analyses and 2D imaging, found no evidence of lymphatic vessels, at least in the scenarios the 2023 paper focused on. But they did detect tubes invading bone in mice with a condition mimicking Gorham-Stout disease. “This [shows] our technology is able to see lymphatic vessels inside the bone” when they are present, Zhou explains.
Several scientists in lymphatics research find much of the critique convincing. Ben Hogan, a developmental biologist at the University of Melbourne and the Peter MacCallum Cancer Centre, says if there really are lymphatic vessels within healthy bone, as Kusumbe’s team argues, it shouldn’t be so hard to find them.
The critique is “careful and rigorous,” adds Jian-Fu Chen, a craniofacial biologist at the University of Southern California who previously examined the skulls of mice and documented lymphatic vessels only on the outer surface of bones.
Kusumbe, now at Nanyang Technological University Singapore, strenuously rejects the critics’ conclusions. Her group’s rebuttal, which includes four of the 11 authors on the 2023 paper, argues the critique doesn’t properly replicate their experiments, and suffers from its own technical problems, including imaging issues that hinder the detection of lymphatic vessels. She argues that other groups’ recent findings are consistent with her lab’s work. And earlier this month, her team posted a preprint underlining the vessels’ role in bone regeneration. The question of the tubes’ existence in bone “should be considered to be resolved,” she says.
The dispute has proved something of a headache for Cell. The journal had originally planned to publish the two papers on 15 July. But it backtracked with a day to spare after Adams raised concerns about the rebuttal, which was first shared with him by Science. In the original version, Kusumbe and colleagues detailed broad criticisms of Adams’s work, including claiming errors in some of his previous papers on blood vessels. The version published this week underwent substantial edits to remove those and other contentions.
Kusumbe, who has previously clashed with Adams online over the two groups’ research findings, says Cell asked her to remove criticisms of Adams’s papers—a decision she found concerning as the rebuttal had already been peer reviewed. (Her team has since posted some of those criticisms as a preprint; Adams says he stands by his research.)
Cell declined to answer questions about its decision-making, saying in a statement: “Because of our responsibility to uphold the confidentiality of the peer-review process, we are unable to elaborate on the details of our correspondence with authors, how this situation might compare with others that may have occurred in our publication history, or any impact it might have on policy.”
Other researchers who saw both versions of the rebuttal tell Science they believe the journal made the right decision to delay publication. “A more focused response specifically addressing the presence of lymphatics in healthy bone [was] a reasonable thing to ask for,” Hogan says.
The debate on these vessels is nevertheless likely to continue, he adds. “It’s easier to suggest that something’s present … than it is to prove that something is absolutely not there.”
Important questions remain about lymphatics’ role in supporting bone, wherever the vessels are situated, Birdsey stresses. “Maybe this will encourage new techniques, new technology to try to answer these criticisms.”