Day: 06/01/2022

Patients with fish allergies may also react to crocodile meat

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Individuals with fish allergies may be at risk for an allergic reaction to crocodile meat and should consult a specialist before consuming it, according to a study published in Pediatric Allergy and Immunology.

As part of their search for alternatives to fish for allergic individuals, the researchers traced this risk to the major fish allergen parvalbumin (PV) present in crocodile meat after observing a pair of anaphylaxis cases involving children with chicken and fish allergies related to PV who ate crocodile meat.

Crocodile

“It turned out that, to promote a healthy and safe diet, we had to also evaluate vertebrates other than bony fish for their potential to trigger an allergic reaction,” Andreas L. Lopata, PhD, associate professor and lead of the molecular allergy research laboratory at the Australian Institute of Tropical Health and Medicine, James Cook University, Townsville, Queensland, Australia, told Healio.

The study is part of a larger project on fish allergies involving more than 100 children from The Children’s Hospital at Westmead in Sydney, funded by the Australian Research Council and National Health and Medical Research Council, aiming for improved diagnostics and management, Lopata said.

Andreas L. Lopata

“Crocodile meat is eaten worldwide by both children and adults. However, the published reports of anaphylaxis were in children who tried crocodile as an alternative to fish/chicken, which they had to avoid. Interestingly, Australia accounts for 60% of the global trade of crocodile products, including meat,” Lopata said.

Alpha PV, which can be found in cartilaginous fish such as shark and ray, is less allergenic than beta PV, which can be found in bony fish such as seabass and salmon. According to the researchers, up to 95% of individuals with fish allergies demonstrate IgE binding to beta PV in bony fish muscle. Crocodiles, the researchers noted, evolved from early bony fish.

Aiming to characterize the IgE binding proteins in crocodile meat, the researchers collected muscle tissue from saltwater crocodile, Asian seabass, Atlantic salmon, ghost shark and bluespotted stingray.

The researchers then recruited 77 children who had a history of IgE-mediated symptoms after eating fish. Skin prick testing to tuna, salmon and seabass produced positive results in 97% of these patients.

Next, eight of 12 of the subjects who were allergic to fish and who had SPT testing to crocodile demonstrated in vivo skin reactivity. By comparison, four individuals with shellfish allergy who received crocodile SPT showed no skin reactivity.

The researchers conducted analyses using commercial and in-house antibodies in addition to individual and pooled patients’ serum to identify and quantify IgE binding proteins, finding that 44 patients (57%) had IgE binding to at least one PV isoform.

Specifically, 31 (70%) showed IgE binding to crocodile PV, 13 (30%) showed IgE binding to both alpha and beta PV, 14 (32%) showed IgE binding only to beta PV and four (9%) showed IgE binding only to alpha PV. Plus, according to the researchers, the signal to beta PV was up to 500 times stronger than the signal to alpha PV.

Although alpha PV was the most abundant isoform, the researchers found that beta PV was more frequently and more strongly recognized by IgE antibodies likely because of its greater similarity including sequence identity to beta PVs in fish, making it the major IgE-binding allergen in bony fish and crocodile.

“The major crocodile protein allergen, parvalbumin, has a higher IgE-antibody binding capacity than some fish parvalbumin. Generally, most cartilaginous fish (sharks and rays) and some bony fish appear much safer than crocodile for fish-allergic patients,” Lopata said.

The researchers assigned Cro p 1 to crocodile beta PV and Cro p 2 to crocodile alpha PV as allergen names and registered them with WHO and the International Union of Immunological Societies.

“Crocodiles are much more dangerous than we ever thought, even when dead on the plate,” Lopata said.

Lopata cautioned patients with fish allergies to avoid consuming alligator and crocodile meat unless medical providers have confirmed their tolerance.

“Fish allergy is highly complex with over a thousand different species consumed worldwide. Because of limited diagnostic capacity, it can be best to recommend the avoidance of all fish and fish products. However, the allergy is often species-specific and can extend to meat from other vertebrates such as crocodiles,” Lopata said.

Also, Lopata said, doctors should be aware of the fish-crocodile syndrome, be cautious about recommending alternatives to fish for allergic individuals and advocate for the development of better diagnostic and management tools.

The researchers also called for further research to improve the accuracy of determining the clinical relevance of novel allergens, including the development of corresponding blood diagnostics and management tools, which could prevent the need for oral food challenges. Currently, the researchers are exploring alternative menus for people with food allergies as well.

For more information:

Andreas L. Lopata, PhD, can be reached at andreas.lopata@jcu.edu.au.

PERSPECTIVE

BACK TO TOP Jennifer Bufford, MS, CRQM, CCRP, CCRC)

Jennifer Bufford, MS, CRQM, CCRP, CCRC

Cross-reactivity in allergic reactions occurs when the proteins in one substance are similar to the proteins found in another substance. As a result, the immune system sees them as the same. In the case of food allergies, cross-reactivity can occur between one food and another. More than 70% identity in the primary sequence is generally needed for cross-reactivity.

Examples of food families with high rates of cross-reactivity include mammalian milks, eggs, fish, shellfish and certain tree nuts. Low rates are noted for grains such as wheat, barley and rye, and rates of cross-reactivity are variable for most other foods.

Cross-reactivity can also happen between pollen and foods or latex and foods. For example, if you are allergic to birch tree pollen, you may also find that eating apples causes a reaction for you.

Also, positive SPT responses to multiple fish in patients with fish allergy is almost the rule, and clinical cross-reactivity is common.

If a person has a measurable reaction to a certain food with clinical diagnosis, avoiding similar foods that could trigger this reaction may be helpful. Fish-allergic individuals, then, should consult with their allergist before consuming crocodilian meat.

This research resulted in the identification of the major allergenic protein, which could help investigators develop a purified extract to aid diagnosis. Such a purified extract could potentially be employed for oral, sublingual or epicutaneous immunotherapy.

Cross-reactivity between fish and crocodile could further be investigated by conducting a study that uses oral food challenges, which are the gold standard for diagnosing a food allergy. Relying solely on a positive IgE response or SPT is insufficient to definitively conclude the presence of a food allergy. The only reliable measure is a failed OFC. In the absence of such data, the authors may be overestimating the prevalence of allergy to crocodile meat.

Jennifer Bufford, MS, CRQM, CCRP, CCRC

Vice President of Clinical Operations, Food Allergy Research & Education

Q&A: U.S. Task Force considers routine chronic kidney disease screening

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The United States Preventive Services Task Force added screening for chronic kidney disease to its list of preventive services under active consideration, according to a press release.

Organizations like the National Kidney Foundation (NKF) and the Coalition for Kidney Health have advocated for the approval of CKD screening since the last time the United States Preventive Services Task Force (USPSTF) considered it in 2012. In 2020, the two groups nominated CKD for another evaluation, referencing new medications and evidence. Nearly 2 years later, the USPSTF has added kidney disease to the list of services it will consider.

"There's a series of new medications that show great promise to slow progression of kidney disease, but these can only work if we're able to identify the people who can benefit from these." Sylvia E. Rosas, MD, MSCE
Sylvia E. Rosas, MD, MSCE, nephrologist and epidemiologist at the Joslin Diabetes Center in Boston and president-elect of the NKF.

Healio sat down with Sylvia E. Rosas, MD, MSCE, nephrologist and epidemiologist at the Joslin Diabetes Center in Boston and president-elect of the NKF, to further discuss the possibility of routine CKD screenings.

Healio: What do you think has changed between now and the last time the USPSTF considered CKD screening in 2012?

Rosas: In the last 5 years or so, we’ve seen an increasing number of FDA approved medications for patients with diabetic kidney disease and without diabetes, that have slowed progression of kidney disease.

The last time we had a positive trial to slow progression to kidney failure in diabetes was for medications called ARBs, which stands for angiotensin receptor blockers, and that was in 2001. Now, we have a variety of new medications. One class is called SGLT2 inhibitors. We also have, more recently, another class of medications called nonsteroidal mineralocorticoid receptors. In addition to that, we have other promising medications that have been shown to slow or decrease the amount of protein that is in urine. Those are called GLP-1 inhibitors and are now under study to see if they slow progression to kidney failure too.

So, there’s a series of new medications that show great promise to slow progression of kidney disease, but these can only work if we’re able to identify the people who can benefit from these therapies.

Healio: The approval of a screening can take up to 3 years. What does that timeline look like?

Rosas: The task force follows guidelines when it comes to their approval process. It will take some time for them to convene the experts, then they will reach out to another group of individuals to review the literature and create a summary. Then they will post for comments and wait to hear from different stakeholders as well as request for evidence that has not been considered. They have a certain series of steps, and the process is transparent. I understand why the process takes longer than we would like because we don’t want them to recommend something that is not based on evidence.

Healio: What obstacles, if any, are in the way of the screening being approved?

Rosas: The task force may be considering other things for screening approval. The hierarchy of those items could potentially push CKD screening for high-risk individuals to the back of the list.

Healio: The Coalition of Kidney Health recently nominated CKD screening to the task force for consideration. Can you talk a bit about NKF’s role in that?

Rosas: Securing a USPSTF screening recommendation has been a long-time priority for the NKF but it has taken on renewed urgency, given the recent work they’ve done to eliminate race as a consideration in the diagnosis of kidney disease. As you know, in 2021, the NKF and ASN joint taskforce issued a report that recommends the adoption of the eGFR 2021 CKD-EPI Creatinine Equation that estimates kidney function without a race variable. NKF has been working tirelessly to facilitate its adoption in the nephrology community and beyond. However, a race-free algorithm is of limited value if people don’t get screened in the first place. We must do more to ensure that at-risk populations get screened and tested as part of a larger strategy to ensure high quality, equitable kidney care.

Healio: What would the approval of CKD screening by the USPSTF mean?

Rosas: CKD screening approval would raise awareness of kidney disease. There is a certain inertia in management because people have this erroneous belief that there’s nothing to be done about kidney disease. It is possible that some people that are at high risk will progress to kidney failure, but we will never know how many kidneys we can save if we don’t start screening early; if we don’t give patients medications; if we don’t treat kidney disease appropriately. We will fail if we don’t try.

So having an indication that USPSTF may approve CKD screening guidelines is fantastic because then we can identify our patients, give them appropriate management, treat their disease early and hopefully prevent people from needing dialysis or kidney transplantation.

Healio: Is there anything you would like to add?

Rosas: Screening for kidney disease involves both a urine test to check for albumin and blood test for creatinine. Patients at high risk for chronic kidney disease, such as individuals with diabetes, hypertension or a family history of kidney disease, should be monitoring their kidney health with these tests. Please visit our website at www.kidney.org if you are interested in learning more about kidney health.

References:

Letter of nomination. https://www.kidney.org/sites/default/files/2020-12-23_ckd_nomination_cover_letter_final.pdf Published Dec. 23, 2020. Accessed May 26, 2022.

Response letter. https://www.kidneynews.org/fileasset/KN/news/uspstf-CKD-nom-acceptance.pdf. Published Feb. 14, 2022. Accessed May 26, 2022.

How to prevent, diagnose and treat monkeypox

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Clinicians should prepare now for the possibility of having to diagnose and treat cases of monkeypox, according to an opinion paper published in Annals of Internal Medicine.

“Physicians should be communicating [to patients] that if they have high-risk sexual contacts and have known contact with somebody who may have had monkeypox-like lesions, they should seek medical attention,” Amesh Adalja, MD, a senior scholar at the Johns Hopkins Center for Health Security and an adjunct assistant professor at the Johns Hopkins Bloomberg School of Public Health, told Healio.

Monkeypox rash. Source: Adobe Stock.
Monkeypox rash. 

Specifically, physicians in primary care, urgent care, emergency medicine, dermatology and sexually transmitted infection (STI) clinics “may be most likely to identify new patients with monkeypox should they continue to appear,” Adalja and Tom Inglesby, MD, the director of the Johns Hopkins Center for Health Security at the Bloomberg School of Public Health, wrote.

Monkeypox virus is in the same orthopoxvirus family as smallpox. The virus had not been detected outside of Africa before 2003, according to Adalja and Inglesby.

Amesh A. Adalja, MD

Amesh Adalja

According to the Center for Infectious Disease Research and Policy, at least 226 cases of monkeypox have recently been identified in 21 countries, including the U.S., Canada and countries in Europe. The in the U.S.

“What distinguishes these cases — all of which are outside the endemic region of the virus — is that person-to-person transmission is occurring, with the majority of cases seemingly unlinked to travel from an endemic country and the appearance of multiple, as-yet unlinked clusters,” Adalja and Inglesby wrote. “This suggests that undetected chains of transmission have been occurring.”

Spread

While the CDC noted that there is a “theoretical risk” for airborne transmission of monkeypox, it is mainly spread from close contact with the skin or fluid of an infected person. In the past, the most consistent source of transmission was through contact with animals, according to Adalja and Inglesby. More recent cases seem to be congregated among men who have sex with men (MSM), and many new infections are being diagnosed at STI clinics, the authors reported.

Incubation of the virus ranges from 5 to 21 days, with a contagious risk occurring when symptoms begin. The mortality risk from infection ranges from 1% to 10%, depending on the availability of medical resources.

Diagnosis

Patients with new onset of febrile illness and rash should be evaluated for monkeypox, according to Adalja and Inglesby. The presence of lymphadenopathy is also a warning sign.

The top clues to watch out for with monkeypox are rash, fever and swollen lymph nodes. Attention should be paid to lesions as well as patients’ social histories due to the disproportionate prevalence of monkeypox among MSM, Adalja said.

Rashes usually start in the mouth before moving to the face and body “in a centrifugal pattern,” Adalja and Inglesby wrote. A PCR test of skin lesions or fluid can confirm a diagnosis. A high suspicion of infection warrants testing.

Treatment

Adalja and Inglesby reported that smallpox antivirals with poxvirus activity such as Vistide (cidofovir, Gilead Sciences), Tembexa (brincidofovir, Chimerix) and TPOXX (tecovirimat, SIGA Technologies Inc) may be used against monkeypox. No standard-of-care treatment for the virus currently exists and only anecdotal reports suggest benefit with antivirals.

“Most patients are not going to have severe enough infection to merit antiviral treatment, but it will be important to study the use of antivirals during this outbreak,” Adalja said.

Researchers behind a British retrospective observational study recently published in The Lancet Infectious Diseases evaluated the use of off-label antivirals administered to seven patients with monkeypox that were detected from 2018 to 2021. Three patients were treated with 200 mg of brincidofovir once weekly; all three patients developed elevated liver enzymes and were forced to suspend therapy. Meanwhile, one patient was treated with 200 mg of tecovirimat twice daily for 2 weeks. This patient experienced no adverse effects and had a shorter duration of viral shedding and illness.

The cohort in this study was too small to conclude any benefit or harm from antivirals for “this neglected tropical disease,” Hugh Adler, MB, BCh, BAO, MRCPI, DTM&H, of the Liverpool School of Tropical Medicine, and colleagues reported.

Overall, “the disease course of the patients … were challenging and resource-intensive to manage, even in the high-income setting of the U.K.,” they wrote.

Prevention

The smallpox vaccine Jynneos (Bavarian Nordic) has an FDA indication for the prevention of monkeypox, and an older-generation vaccine (ACAM2000, Sanofi Pasteur Biologics Co.) may be used off-label for monkeypox, according to Adalja and Inglesby. Clinicians who administer either vaccine immediately after a suspected exposure can “abort infection or significantly attenuate it,” they wrote. For patients who cannot receive the smallpox vaccines, vaccinia immune globulin may be administered.

While researchers attempt to uncover what is driving this outbreak of monkeypox, clinicians should apply “enhanced case finding, isolation, contact tracing and postexposure vaccination,” the authors advised. Any new cases of monkeypox should be reported to a state or local health department immediately.

References:

Adalja A, Inglesby T. Ann Intern Med. 2022;doi:10.7326/M22-1581.

Adler H, et al. Lancet Infect Dis. 2022;doi:10.1016/S1473-3099(22)00228-6.

Multi-country monkeypox outbreak in non-endemic countries. https://www.who.int/emergencies/disease-outbreak-news/item/2022-DON385. Published May 21, 2022. Accessed May 25, 2022.

Officials confirm 226 monkeypox cases in 21 countries. https://www.cidrap.umn.edu/news-perspective/2022/05/officials-confirm-226-monkeypox-cases-21-countries. Published May 25, 2022. Accessed May 26, 2022.

Mars’ emitted energy and seasonal energy imbalance

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Significance

The radiant energy budget is a fundamental metric for planets. Based on the observations from multiple missions, we provide a global picture of Mars’ emitted power. Furthermore, we estimate the radiant energy budget of Mars, which suggests that there are energy imbalances at the time scale of Mars’ seasons. Such energy imbalances provide a new perspective to understanding the generating mechanism of dust storms. Mars’ radiant energy budget is assumed to be balanced at all time scales in current models and theories, but our analyses show that the energy budget is not balanced, at least at the time scale of Mars’ seasons. Therefore, current theories and models should be revisited with the newly revealed energy characteristics.

Abstract

The radiant energy budget of a planet is essential to understanding its surface and atmospheric processes. Here, we report systematic measurements of Mars’ emitted power, which are used to estimate the radiant energy budget of the red planet. Based on the observations from Mars Global Surveyor, Curiosity, and InSight, our measurements suggest that Mars’ global-average emitted power is 111.7 ± 2.4 Wm−2. More importantly, our measurements reveal strong seasonal and diurnal variations of Mars’ emitted power. The strong seasonal variations further suggest an energy imbalance at the time scale of Mars’ seasons (e.g., ∼15.3% of the emitted power in the Northern autumn for the Southern Hemisphere), which could play an important role in generating dust storms on Mars. We also find the 2001 global dust storm decreased the global-average emitted power by ∼22% during daytime but increased the global-average emitted power by ∼29% at nighttime. This suggests that global dust storms play a significant role in Mars’ radiant energy budget.

Current recommendations for cancer surveillance in Gorlin syndrome: a report from the SIOPE host genome working group (SIOPE HGWG)

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Abstract

Gorlin syndrome (MIM 109,400), a cancer predisposition syndrome related to a constitutional pathogenic variation (PV) of a gene in the Sonic Hedgehog pathway (PTCH1 or SUFU), is associated with a broad spectrum of benign and malignant tumors. Basal cell carcinomas (BCC), odontogenic keratocysts and medulloblastomas are the main tumor types encountered, but meningiomas, ovarian or cardiac fibromas and sarcomas have also been described. The clinical features and tumor risks are different depending on the causative gene. Due to the rarity of this condition, there is little data on phenotype-genotype correlations. This report summarizes genotype-based recommendations for screening patients with PTCH1 and SUFU-related Gorlin syndrome, discussed during a workshop of the Host Genome Working Group of the European branch of the International Society of Pediatric Oncology (SIOPE HGWG) held in January 2020. In order to allow early detection of BCC, dermatologic examination should start at age 10 in PTCH1, and at age 20 in SUFU PV carriers. Odontogenic keratocyst screening, based on odontologic examination, should begin at age 2 with annual orthopantogram beginning around age 8 for PTCH1 PV carriers only. For medulloblastomas, repeated brain MRI from birth to 5 years should be proposed for SUFU PV carriers only. Brain MRI for meningiomas and pelvic ultrasound for ovarian fibromas should be offered to both PTCH1 and SUFU PV carriers. Follow-up of patients treated with radiotherapy should be prolonged and thorough because of the risk of secondary malignancies. Prospective evaluation of evidence of the effectiveness of these surveillance recommendations is required.

Conclusion

GS was described many years ago, but the identification of different underlying molecular genetic characteristics (PTCH1 or SUFU germline variants) has changed the way in which this predisposing syndrome is identified and has modified the phenotypic characteristics of the tumor risks historically described. There are still many uncertainties about the risk of tumors associated with GS. Given the rarity of this syndrome, the estimation of the risks is still not very precise and must be improved by studies on a larger number of patients with analysis taking into account ascertainment bias. Nevertheless, due to the risk of tumor, any patient with GS predisposition syndrome should benefit from a specific genotype-based surveillance program as described in these recommendations. The evaluation of the feasibility and efficacy of these recommendations is necessary.

Downregulated exosomal microRNA-148b-3p in cancer associated fibroblasts enhance chemosensitivity of bladder cancer cells by downregulating the Wnt/β-catenin pathway and upregulating PTEN

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Abstract

Objective

Exosomes derived from cancer-associated fibroblasts (CAFs) are known as important drivers of tumor progression. Previously, microRNA (miR)-148b-3p has been found to be upregulated in bladder cancers as well as in body fluids (blood, urine) of bladder cancer patients. Here, we aimed to explore the role of CAF-derived exosome miR-148b-3p in bladder cancer progression and chemosensitivity.

Methods

Transwell, MTT, flow cytometry and colony formation assays were applied to assess the effects of CAF-derived exosomes on bladder cancer cell metastasis, epithelial-mesenchymal transition (EMT) and chemosensitivity. A dual luciferase reporter assay was employed to evaluate the targeting relationship between miR-148b-3p and PTEN. Gain- and loss- of function assays were conducted to explore the roles of miR-148b-3p and PTEN in the behavior of bladder cancer cells. The role of PTEN in the metastasis, EMT and chemosensitivity of bladder cancer cells was assessed both in vivo and in vitro.

Results

We found that CAF-derived exosomes promoted the metastasis, EMT and drug resistance of bladder cancer cells. We also found that CAF-derived exosomes could directly transport miR-148b-3p into bladder cancer cells. In a xenograft mouse model we found that CAF-derived exosomes increased miR-148b-3p expression levels and promoted tumor proliferation, metastasis and drug resistance. PTEN was validated as a target of miR-148b-3p. Concordantly, we found that PTEN overexpression inhibited EMT, metastasis and chemoresistance in bladder cancer cells, reversing the tumor promoting effects of miR-148b-3p via the Wnt/β-catenin pathway.

Conclusions

Our results suggest that miR-148b-3p downregulation in CAF-derived exosomes, thereby inhibiting the Wnt/β-catenin pathway and promoting PTEN expression, may offer potential opportunities for bladder cancer treatment.

Discussion

Resistance to chemotherapy is a frequent cause of tumor recurrence and treatment failure in bladder cancer [25]. A recent study revealed that inhibition of cancer-derived exosomes may increase the sensitivity of bladder cancer cells to cisplatin [26]. This finding inspired us to test the efficacy of bladder cancer treatment from the perspective of exosomes and drug sensitivity. Using a series of assays we were able to confirm our aforementioned hypothesis that a regulatory loop involving miR-148b-3p, PTEN and the Wnt/β-catenin pathway may be active in bladder cancer cells. From our results we conclude that miR-148b-3p silencing in CAF-exosomes may reinforce bladder cancer cell chemosensitivity through inactivating the Wnt/β-catenin pathway via PTEN upregulation.

First, we found that exosomes secreted by CAFs reduced the apoptosis and promoted the metastasis and chemoresistance of bladder cancer cells by enhancing EMT, indicated by higher levels of N-cadherin and vimentin and lower levels of E-cadherin expression. Others have shown that CAFs can secrete FAP and α-SMA, thereby creating a favorable environment for cancer cells and enabling the migration and invasion of cancer cells [27]. In addition, it has been found that exosomes derived from cancer cells have the potential to expel anticancer drugs and induce chemotherapeutic resistance in malignant cells [28]. It has also been found that bladder cancer cell-derived exosomes can induce cell proliferation and inhibit tumor cell apoptosis in vitro in conjunction with upregulated expression levels of Bcl-2, but reduced expression levels of Bax and caspase-3 [29], which is in line with our current observations. Vimentin is a well-recognized metastasis marker [30], while E-cadherin is known to repress the invasion and metastasis of epithelial cells [31]. Additionally, CAF-secreted exosomes have been found to reinforce metastasis and chemoresistance by enhancing EMT in colorectal cancer cells [7]. Based on this information, we propose that inhibition of exosome formation and release may serve as a promising strategy for the treatment of bladder cancer.

Others have shown that abnormal miR-148b-3p levels may be present in sera of bladder cancer patients and, as such, may serve as diagnostic markers [12]. Inspired by this finding, we decided to assess miR-148b-3p expression in CAF-secreted exosomes in order to get insight in its mode of action in bladder cancer. We found that the miR-148b-3p levels were upregulated in bladder cancer cells and CAF-exos and that downregulation of miR-148b-3p in CAF-exos could inhibit EMT, metastasis and drug resistance in bladder cancer cells. It has also been shown that exosomes released from malignant cells may have specific biological activities leading to enhanced proliferation, tumor development, metastasis and resistance to therapy [32]. miR-148b-3p has been shown to strongly correlate with exosome content in paroxysmal nocturnal hemoglobinuria [33]. Similarly, exosomal miR-148a has been found to be overexpressed in the sera of osteosarcoma patients showing a poor chemotherapeutic response [34]. Importantly, it has been found that CAF-secreted exosomal miR-146a may facilitate bladder cancer development [35]. Based on these findings, we speculate that knocking down miR-148b-3p expression in CAF-secreted exosomes may serve as a tool to treat bladder cancer.

We also found that PTEN serves as a target of exosome-mediated miR-148b-3p, and that PTEN upregulation inhibits EMT, metastasis and chemoresistance of bladder cancer cells via the Wnt/β-catenin pathway. PTEN-deficient cells have previously been found to exhibit resistance to rapamycin in advanced transitional cell carcinoma, the most frequently diagnosed type of bladder cancer [36], suggesting a protective role of PTEN in bladder cancer. PTEN deficiency in T24 and HT-1376 bladder cancer cells noticeably attenuated the sh-Jumonji AT-rich interactive domain 2-mediated increase of E-cadherin and decrease of vimentin and N-cadherin expression [37]. Conversely, overexpression of PTEN has been found to inhibit miR-495-mediated proliferation and invasion of bladder cancer cells [38]. Convincing evidence indicates that PTEN loss is associated with urothelial tumor induction, the progression of bladder cancer and its resistance to chemotherapy [39]. A recent study has revealed involvement of PTEN and the Wnt/β-catenin pathway in drug resistance [24], and it has been found that PTEN overexpression can block β-catenin-induced urothelial proliferation and tumorigenesis [18]. In addition, Cullin 4B has been found to promote EMT and metastasis by upregulating the Wnt/β-catenin pathway in bladder cancer [40]. Taken together, we conclude that overexpression of PTEN may reverse the tumorigenic effects of exosomal miR-148b-3p in bladder cancer.

In summary, we found that CAF-exos and exosomal miR-148b-3p can reduce apoptosis and promote EMT, metastasis and drug resistance in bladder cancer cells and that these effects can be reversed by PTEN overexpression via downregulation of the Wnt/β-catenin pathway. These results may be relevant for future (pre-)clinical investigations and applications.

A novel lymphatic pattern promotes metastasis of cervical cancer in a hypoxic tumour-associated macrophage-dependent manner

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Abstract

Lymphatic remodelling in the hypoxic tumour microenvironment (TME) is critically involved in the metastasis of cervical squamous cell carcinoma (CSCC); however, its underlying mechanisms remain unclear. Here, we uncovered a novel lymphatic pattern in the hypoxic TME, wherein lymphatic vessels (LVs) are encapsulated by tumour-associated macrophages (TAMs) to form an interconnected network. We describe these aggregates as LVEM (LVs encapsulated by TAMs) considering their advantageous metastatic capacity and active involvement in early lymph node metastasis (LNM). Mechanistic investigations revealed that interleukin-10 (IL-10) derived from hypoxic TAMs adjacent to LVs was a prerequisite for lymphangiogenesis and LVEM formation through its induction of Sp1 upregulation in lymphatic endothelial cells (LECs). Interestingly, Sp1high LECs promoted the transactivation of C–C motif chemokine ligand 1 (CCL1) to facilitate TAM and tumour cell recruitment, thereby forming a positive feedback loop to strengthen the LVEM formation. Knockdown of Sp1 or blockage of CCL1 abrogated LVEM and consequently attenuated LNM. Notably, CSCCnon-LNM is largely devoid of hypoxic TAMs and the resultant LVEM, which might explain its metastatic delay. These findings identify a novel and efficient metastasis-promoting lymphatic pattern in the hypoxic TME, which might provide new targets for anti-metastasis therapy and prognostic assessment.

Discussion

Tumour heterogeneity in the hypoxic TME is a potent driver of tumour progression and metastasis [5]. LVs in the hypoxic TME show remarkable plasticity and heterogeneity, reflecting their functional specialization [6]. Therefore, therapeutic strategies that non-selectively target the entire LV population are ineffective and can potentially lead to cancer progression [29]. In the present study, we discovered a novel lymphatic pattern and determined its clinical significance in CSCC. We found that the LVEM in hypoxic milieu provided an important pathway for lymphatic metastasis, by which activated LECs recruited tumour cells in a hypoxic TAM-dependent manner. Interestingly, CSCCnon-LNM is largely devoid of this LVEM, which might explain its metastatic delay and lower potential for metastasis. Moreover, the highly aggressive LVEM appears to be essential to tumour cell dissemination and might substantially contribute to the biological differences between CSCCLNM and CSCCnon-LNM tissues. Thus, we provided mechanistic and clinical insight into the role of the LVEM in lymphatic metastasis.

In our previous study, we reported preferential accumulation of TAMs in hypoxic areas of tumours and their secretion of cytokines promoting the formation of pro-metastatic tumour niches [23]. Additionally, we found that in the hypoxic zones of a primary tumour site, a subset of TAMs adjacent to LVs constructed a novel lymphatic metastasis-promoting pattern. The general understanding of how TAMs influence lymphangiogenesis focuses on their production and release of growth factors and metalloproteases [30]. Nevertheless, the mechanisms by which a subset of TAMs adjacent to LVs participate in the formation of lymphatic conduits within the hypoxic TME, as well as their cooperation to facilitate tumour cell intravasation into LVs, remain unidentified. A recently identified mechanism suggests that podoplanin-expressing TAMs engage β1 integrins during the recruitment and adhesion of these cells to galectin-8-expressing lymphatics [31]. Once in the perilymphatic space, TAMs promote endothelial cell remodelling to enhance lymphangiogenesis and metastasis. In the present study, we identified and characterized a subset of hypoxic TAMs that localized proximal to tumour lymphatics and were relevant to lymphoinvasion. Both findings emphasize a TAM subset that specifically interacts with LVs to promote neo-lymphatic formation and the migration of tumour cells through the lymphatic system. Our study highlights the unique aggressive nature of the lymphatic metastasis-promoting pattern, which might emerge as a major characteristic to discriminate between CSCCLNM and CSCCnon-LNM patients.

Hypoxic TAMs form a favourable niche for LVEM formation through IL-10 upregulation. CM collected from LECs treated with IL-10 stimulated tumour cell chemotaxis to an even greater extent than CM obtained from untreated LECs. Our data revealed that hypoxic TAMs but not tumour cells were the predominant source of IL-10. IL-10 stimulation also leads to a further upregulation of IL-10 in macrophages, which is an autocrine amplification cascade incited by IL-10 [32]. IL-10 is a multifunctional cytokine involved in anti-inflammatory and immunoregulatory effects [33]. In addition to its wide-ranging immunomodulatory functions, IL-10 also regulates lymphangiogenesis via macrophages [34]. In the present study, our data indicated that IL-10 was a potential mediator of cross-talk between TAMs and LECs in hypoxic zones, with this activity involved in mediating LVEM formation. In these zones, IL-10 is also an important determinant of alternative M2 activation and sustainment of tumour cell proliferation [35]. Higher lymphatic activity is mainly regarded as an increased opportunity for tumour cells to access the lymphatic system [36]. Blockade of IL-10 signalling resulted in abrogation of TAM-H-mediated LEC activation, and the observed increase in tumour cell chemotaxis to inflamed LECs was completely abolished by depleting IL-10R. However, IL-10 did not have a direct effect on tumour cell migration. Previous reports indicate that CSCC patients exhibiting upregulated levels of serum IL-10 showed poorer response to antitumour therapy relative to patients with low serum IL-10 levels [37]. The results of our present study revealed that hypoxic TAMs exhibiting elevated IL-10 expression formed a favourable niche for lymphangiogenesis and LVEM formation and promoted metastatic progression of CSCC, which might explain the poor therapeutic response of these patients.

We then examined the regulatory mechanisms of IL-10-mediated LVEM in CSCCLNM tissues, finding that the niches formed by IL-10-activated LECs provided a constant source of paracrine CCL1 for TAM infiltration, which in turn further strengthened the LVEM. Chemokines were recently implicated in organ-specific metastasis of tumour cells [38], and human CCL1 is a potent monocyte chemoattractant that binds to and exclusively activates CCR8 [39]. In the present study, adding an anti-CCL1 neutralization antibody to IL-10-activated LEC CM or silencing CCR8 on receptor cells using siRNA or a CCR8 antagonist significantly impaired cell migration, indicating that the CCL1–CCR8 axis mediates most of the chemotactic response to LEC CM. CCR8 plays a rather unique role in regulating the immune response [40] and is preferentially expressed by activated T helper-type 2 cells and tumour cells [15], which mediates their recruitment to the lymphatics. Moreover, the CCL1–CCR8 axis also controls entry of other tumour-infiltrating lymphocytes into LNs for microenvironment remodelling [41]. For tumours to metastasize, tumour cells must gain enhanced migratory capacity, and the TME must be remodelled [42]. CCL1 produced by LECs mediates TAM migration towards LECs during LVEM formation while also promoting tumour cell entry into LNs by increasing tumour cell motility [43]. These results suggest that metastatic tumour cells are responsive to the chemotactic signals from lymphatic endothelium. Moreover, chemokines derived from the endothelium are essential for integrin-mediated adhesion and transendothelial cell migration [44]. These data not only emphasize the indispensable role of LVEM in LNM but also indicate that CCL1 might represent an attractive alternative therapeutic target to interfere with CSCC metastasis.

In this scenario, there is a pressing need to identify more specific and convenient markers to distinguish metastasis-associated LVs for precision treatment. Our analysis of the CCL1 promoter region identified Sp1 as a transcription regulator in LECs that contributes to CSCC metastasis. Furthermore, LVEMhigh CSCC tissues displayed a higher Sp1 and CCL1 level relative to that in LVEMlow cases. Sp1 is a highly regulated transcription factor involved in regulating a large number of genes that contribute to the “hallmarks of cancer” [45]. Sp1 also involved in upregulating the lymphangiogenic genes, such as VEGF-C, VEGFR-3 to promote lymphangiogenesis [46, 47]. Additionally, Sp1 is essential for early embryonic development but dispensable for cell growth and differentiation [48], with Sp1 expression changing during development and varying in different cell types [49]. Previous studies reported that Sp1 expression correlated with dismal patient outcome in various cancer types, including CSCC [50]. Notably, accumulating evidence shows that Sp1 plays a critical role in the inflammatory signalling that mediates cancer-stroma cross-talk [51]. Therefore, investigating the actions associated with aberrant activation of Sp1 is important to understanding tumour progression. In this study, Sp1 was suggested as a downstream TF of the IL-10/STAT3 pathway. Moreover, we found that metastasis-associated LVs could play an active role in tumour metastasis, which was at least partly due to elevated expression of Sp1. Importantly, we confirmed that blockade of Sp1 or CCL1 prevented TAM proximity to LVs, thereby reducing tumour lymphangiogenesis and metastatic dissemination. This suggests that identification of the Sp1high LVEM subset would have a profound impact on prognosis assessment. Our data revealed Sp1 as a key factor in a previously unexplored metastatic cascade in CSCC and promote further screening and potential development of Sp1-specific inhibitors in cancer therapy. Furthermore, analysis of Sp1high LVs in a tumour could be an indication for complete LN resection or adjuvant therapy. The prognostic values of LVs previously denoted by conventional markers, such as LYVE-1 or D2-40, are often different or even opposing in different studies [52, 53]. Indeed, single expression of LYVE-1 cannot precisely predict LNM states [54]. The present study provides a molecular definition for metastasis-associated LVs that suggests anti-Sp1 therapy as potentially beneficial for LVEMhigh CSCC patients through its inhibition of LVEM formation. A larger cohort of patients is warranted to further explore this idea.

Lymphatic metastasis is the principal reason for the poor survival rates of CSCC patients. A detailed understanding of how LVs contribute to further metastasis is crucial to better comprehend the biological complexity of lymphatic metastasis. In this study, we defined a lymphatic metastasis-promoting pattern characterized by a lymphatic skeleton surrounded by TAMs, with this pattern prevalent in CSCCLNM tissue and actively involved in lymphatic metastasis. Moreover, M2 macrophages were selectively recruited by Sp1high LECs to form this unique metastasis-promoting pattern. Identification of the LVEM and its regulatory mechanisms not only offers novel targets for the development of anti-metastasis therapy but also provides a basis for the selection of specific cohorts of patients who might benefit from certain molecular-targeted drugs.

Drinking unsweetened or sweetened coffee linked to lower death risk: New Study

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A new study has found that in comparison to non-coffee drinkers, adults who drank moderate amounts (1.5 to 3.5 cups per day) of unsweetened coffee or coffee sweetened with sugar were less likely to die during a 7-year follow-up period.

The results for those who used artificial sweeteners were less clear. The findings are published in Annals of Internal Medicine. Previous studies observing the health effects of coffee have found that coffee consumption is associated with a lower risk of death but did not distinguish between unsweetened coffee and coffee consumed with sugar or artificial sweeteners.

Researchers from Southern Medical University in Guangzhou, China used data from the U.K. Biobank study health behaviour questionnaire to evaluate the associations of consumption of sugar-sweetened, artificially sweetened, and unsweetened coffee with all-cause and cause-specific mortality.

More than 171,000 participants from the U.K. without known heart disease or cancer were asked several dietary and health behaviour questions to determine coffee consumption habits. The authors found that during the 7-year follow-up period, participants who drank any amount of unsweetened coffee were 16 to 21 percent less likely to die than participants who did not drink coffee.

They also found that participants who drank 1.5 to 3.5 daily cups of coffee sweetened with sugar were 29 to 31 percent less likely to die than participants who did not drink coffee. The authors noted that adults who drank sugar-sweetened coffee added only about 1 teaspoon of sugar per cup of coffee on average. Results were inconclusive for participants who used artificial sweeteners in their coffee.

Any accompanying editorial by the editors of Annals of Internal Medicine notes that while coffee has qualities that could make health benefits possible, confounding variables including more difficult to measure differences in socioeconomic status, diet, and other lifestyle factors may impact findings. The authors add that the participant data is at least 10 years old and collected from a country where tea is a similarly popular beverage.

They caution that the average amount of daily sugar per cup of coffee recorded in this analysis is much lower than specialty drinks at popular coffee chain restaurants, and many coffee consumers may drink it in place of other beverages that make comparisons to non-drinkers more difficult.

Based on this data, clinicians can tell their patients that there is no need for most coffee drinkers to eliminate the beverage from their diet but to be cautious about higher calorie specialty coffees.

Stop counting calories and focus on whole foods to lose weight and get healthier, says scientist

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Forget about calories and focus on ingredients to lose weight, Dr. Tim Spector told Insider.

Ultra-processed foods can be less satisfying than whole foods, causing us to overeat.

People should eat fewer ultra-processed foods (UPFs) instead of counting calories to lose weight and improve their health, according to epidemiologist Dr. Tim Spector.

A calorie deficit is required for weight loss, but Spector said that calorie tracking is inaccurate because labels are often incorrect, and people will naturally consume fewer calories if they prioritize eating whole foods without any artificial ingredients.

“We’ve got to stop talking about calories,” Spector, professor of genetic epidemiology at King’s College London, UK, said. “Ultra-processed food is the enemy.”

Spector is the author of best-selling book, “Spoon-Fed: Why Almost Everything We’ve Been Told About Food is Wrong.”

Lots of foods are processed in some way, such as oats, yogurt, or flour. But UPFs are generally mass-produced in a factory and contain additives such as colors and preservatives to maintain taste or texture, which have been linked to various health problems. Examples of UPFs include packaged cakes and cookies, and bags of chips.

Research suggests that more than half of the foods consumed in high income countries are UPFs.

A small 2019 study from the National Institutes of Health found that diets high in UPFs led people to eat on average 500 more calories a day and gain more weight when compared with those eating fresh food, as Insider’s Hilary Brueck reported. Scientists aren’t sure why this happened, but they think there may be something unique about how our hormones interact with processed foods.

UPFs have also been linked to people dying earlier and a higher risk of heart disease, according to a 2020 study published in the American Journal of Clinical Nutrition, as Brueck and Insider’s Gabby Landsverk reported.

‘It’s a myth that calories are useful’

Spector believes that the biggest myth damaging people’s health is that calories are useful. Their importance is “vastly overblown” and we should be more concerned with the ingredients in food, he said.

“We all have very different metabolic needs so you can’t rely on how someone else did on some calorie-controlled diet or stick to some totally arbitrary 2,000 calories for women, it’s complete rubbish,” he said.

For example, while a croissant is generally fewer calories than avocado and eggs on sourdough toast, the latter has a higher nutritional value and will keep you fuller for longer due to the fiber, protein, and healthy fats.

Whole foods can keep you fuller than ultra-processed foods

Spector said calorie-counting drives people to eat more UPFs because they will seek out low calorie versions of food.

Brands remove fat and sugar from foods to reduce the number of calories, and sometimes add chemicals and sweeteners to ensure they are still palatable.

Research shows people can end up overeating low-calorie but highly processed foods which aren’t always as satiating, which can lead to weight gain.

Whole foods are more satiating than UPFs because they are usually higher in fiber and protein, so they release energy slowly and keep people full, making them less likely to reach for more food soon after, Spector said.

Nick Shaw, a bodybuilder, personal trainer and diet coach at RP Strength previously told Insider that he encourages people to prioritize whole foods when losing weight, even if less nutritious foods have the same calories and macros.

“The best idea is to use food quality as a means to easier control your food quantity,” he said. “If you stick to mostly whole food choices like lean proteins (eg. chicken breast), lots of vegetables, healthy fats, and then some healthy carbs (eg. sweet potato or whole grains) you will feel more full, have fewer cravings, and you can still hit your food quantities needed to help with fat loss.”

Avoid foods with more than 10 ingredients

While some foods are technically processed, they aren’t ultra-processed and can fit into a healthy diet and don’t need to be avoided. These include canned fruit, frozen vegetables, and yogurt, Spector said.

To work out whether a food is ultra-processed, look at the ingredients list and see how long it is — if a food has more than 10 ingredients, it might not be a great choice.

See how many names you recognize, too — if you see something you wouldn’t have in your kitchen cupboard, it’s best to steer clear, Spector said.

3 pelvic-opening yoga asanas to regulate your menstrual cycle

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Periods, the more common term for menstrual cycle, is a natural process that takes place in a female body in order to get rid of blood and tissues through the uterus. The menstrual cycle is the time between two consecutive periods and it is mostly 28 days but can be anything between 26-32 days. However, most women, due to today’s hectic lifestyle patterns, experience the problem of irregular periods, which means their periods are delayed by more than a week after the due date.

While a healthy diet and constant workout is the best way to regulate periods without any medications or pills, there are certain yoga asanas, as recommended by fitness trainer Juhi Kapoor, which can help improve your condition.

In her Instagram post, she mentioned how these asanas are great for pelvic opening, strengthening the pelvic and improving blood flow to the reproductive area.

Here are the three yoga asanas that you can try for regulating your menstrual cycle

1. Utkata Konasana or Goddess Squat

This yoga asana helps to open the hips and chest while strengthening and toning the lower body. It also stimulates the uro-genital, respiratory and cardiovascular systems for overall betterment of female health.

2. Skandasana or Side Lunges

It enhances your body balance and core strength apart from stretching the hamstrings and hips, which often get due to a sedentary lifestyle. Overall, it also helps to relieve back pain and prevent sciatica.

3. Malasana or Garland Pose

The well-known yoga pose not only improves balance and concentration, and focus but also increases circulation and blood flow in the pelvis. Not just menstruating women, it is particularly good for pregnant women as well.

All the three yoga asanas are highly recommended to treat PCOS and endometriosis, improve period flow, boost fertility in both men and women apart from leg toning and strengthening. These poses also ease the birth canal opening during delivery.

However, do not perform these asanas if you have knee, ankle and heel pain, or Arthritis. Also avoid doing them in the first trimester of pregnancy, and in the second and third trimester, use the necessary support of the wall or chair to maintain balance.