Mois : juillet 2024

Researchers publishing in Probiotics and Antimicrobial Proteins have published the results of a clinical trial on the effects of a probiotic on inflammaging in older people.

The gut and systemic inflammation

The researchers begin this paper discussing their target of inflammaging, as this condition has been linked to multiple other age-related disorders, including Alzheimer’s [1]. Inflammaging has been found to have multiple causes, one of them being a change in gut bacteria [2]. The intestinal bacteria’s gradual transition from a younger to an older phenotype has been well-documented [3], but strategies for re-colonizing the human gut with probiotics have only been somewhat explored, with one trial demonstrating effectiveness against the age-related muscle loss known as sarcopenia [4].

Therefore, these researchers decided to perform a clinical trial of probiotics against inflammaging, with the endpoints being biomarkers of inflammation such as C-reactive protein (CRP). Because some compounds naturally found in berries have been previously found to have antioxidant [5] and anti-inflammatory [6] properties, berries were also included to determine if their effects could be complementary to probiotics.

Largely inconclusive results with bright spots

An active version of Lactiplantibacillus plantarum HEAL9 (LpHEAL9) was chosen as the probiotic for this study. Out of 187 screened subjects, 66 people had enrolled into it, with 22 being allocated to each group: placebo control, LpHEAL9, and LpHEAL9 with berries. The participants were all in their seventies, with a mean age of 73.2.

Unfortunately, this study did not meet its CRP endpoint. While there was a trend towards decreasing CRP in the LpHEAL9-only group, this result did not meet statistical significance, and other results also trended similarly. However, the LpHEAL9-only group had less of the inflammatory biomarker calprotectin in their feces, and this result was statistically significant. This group also appeared to have less of the inflammatory factor TNF-alpha, although this result did not quite reach statistical significance, and there were no differences in other serum biomarkers.

Some positive trends were found in cognition. The LpHEAL9-only group had trends towards better performance on one of the cognitive trail-making tests compared to either of the other groups, and the berries-included group appeared to perform slightly better on the other test; these results, too, did not reach statistical significance.

When examining a variety of other metrics, the researchers found a beneficial, statistically significant result in body pain, which was decreased in the group that included berries. There was also a harmful result: while over the course of the study, the systolic blood pressure of the placebo group was significantly reduced, this occurred to neither of the LpHEAL9 groups. Many other metrics, including most gastrointestinal metrics, did not have any statistically significant results. The bowel function of the LpHEAL9-only group was disrupted for the first three weeks, with an increase in bowel movements, but this returned to normal over time.

While these results are largely disappointing and suggest a lack of efficacy for this probiotic treatment against inflammaging-related conditions, the calprotectin result suggests that there are potential uses for LpHEAL9. In a previous study, a different strain of this bacterium, CJLP243, was also found to reduce calprotectin, and this significantly reduced diarrhea in a population that had elevated levels of this biomarker [7]. Calprotectin is also a key biomarker in determining inflammatory from non-inflammatory bowel syndromes [8].

These largely mixed results suggest that berry supplementation, in addition to probiotics, may be useful in decreasing overall pain, although this result will have to be confirmed with a separate study. While this particular probiotic may not be effective against inflammaging in general, this or other probiotics may be useful in treating inflammatory gut conditions.

Literature

[1] Franceschi, C., Bonafè, M., Valensin, S., Olivieri, F., De Luca, M., Ottaviani, E., & De Benedictis, G. (2000). Inflamm‐aging: an evolutionary perspective on immunosenescence. Annals of the new York Academy of Sciences, 908(1), 244-254.

[2] Li, X., Li, C., Zhang, W., Wang, Y., Qian, P., & Huang, H. (2023). Inflammation and aging: signaling pathways and intervention therapies. Signal Transduction and Targeted Therapy, 8(1), 239.

[3] Mariat, D., Firmesse, O., Levenez, F., Guimarăes, V. D., Sokol, H., Doré, J., … & Furet, J. (2009). The Firmicutes/Bacteroidetes ratio of the human microbiota changes with age. BMC microbiology, 9, 1-6.

[4] Qaisar, R., Burki, A., Karim, A., Iqbal, M. S., & Ahmad, F. (2024). Probiotics supplements Improve the sarcopenia-related quality of life in older adults with age-related muscle decline. Calcified Tissue International, 114(6), 583-591.

[5] McGhie, T. K., Walton, M. C., Barnett, L. E., Vather, R., Martin, H., Au, J., … & Kruger, M. C. (2007). Boysenberry and blackcurrant drinks increased the plasma antioxidant capacity in an elderly population but had little effect on other markers of oxidative stress. Journal of the Science of Food and Agriculture, 87(13), 2519-2527.

[6] Landete, J. M. (2011). Ellagitannins, ellagic acid and their derived metabolites: A review about source, metabolism, functions and health. Food research international, 44(5), 1150-1160.

[7] Jung, M., Jung, S., Kim, N., Ahn, H., Yun, H., & Kim, K. N. (2022). A randomized, double-blind, placebo-controlled trial to assess the efficacy and safety of Lactiplantibacillus plantarum CJLP243 in patients with functional diarrhea and high fecal calprotectin levels. Nutrients, 14(2), 389.

[8] Menees, S. B., Powell, C., Kurlander, J., Goel, A., & Chey, W. D. (2015). A meta-analysis of the utility of C-reactive protein, erythrocyte sedimentation rate, fecal calprotectin, and fecal lactoferrin to exclude inflammatory bowel disease in adults with IBS. Official journal of the American College of Gastroenterology| ACG, 110(3), 444-454.

By making NK cells insensitive to tumor-secreted TGF-β, scientists have improved their efficacy against this deadly hepatocellular carcinoma (HCC) [1].

Liver cancer and TGF-β

Cancer cells, which are supposed to be vulnerable to the immune system, develop various defensive mechanisms to avoid detection and decrease immune cells’ fitness and viability. In solid tumors, the dense tumor microenvironment (TME) has a particularly strong dampening effect on immune activity.

HCC is the most widespread form of liver cancer. It is also very deadly, with a 5-year survival rate of around 20%. Many therapies have been tried and failed against HCC.

HCC is also one of the cancers that is characterized by overexpression of transforming growth factor beta (TGF-β). As its name hints at, TGF-β drives tumor growth, but it also interferes with the activity of immune cells [2], such as natural killer (NK) cells, which are part of the innate immune system.

Natural born killers, improved

In this new study, scientists from the University of California – San Diego combined several cutting-edge techniques to try and overcome this problem. To begin with, they used cellular reprogramming to manufacture brand-new NK cells. This involves producing induced pluripotent stem cells (iPSCs) from differentiated cells and their further re-differentiating them into NK cells.

However, there was a twist: in those iPSCs, genetic changes were introduced to make them resistant to TGF-β, either by knocking out TGF-β receptor 2 (TGFBR2) or by expressing its double-negative version, which also blocks TGF-β signaling. The researchers assumed that such cells would be less susceptible to the inhibitory effect of TGF-β secreted by HCC cells.

Their gamble seemed to pay off: the resulting NK cells showed superior cytotoxicity in an in vitro model of HCC, compared to wild type NK cells. They also retained functional activity when pre-treated with TGF-β, unlike wild type NKs. The only problem was that iPSCs lacking functional TGF-β receptors reproduced slower (since TGF-β boosts growth).

The researchers tried to further improve the efficacy of their modified NK cells by arming them with chimeric antigen receptors (CARs) that are routinely used against HCC. CARs are usually associated with T cells, but they can be introduced to other types of immune cells to direct them towards antigen-expressing cancer cells.

NK cells with TGFB2R knocked out and without CAR expression killed HCC cells better than CAR-expressing wild type NK cells. There was some synergy between CARs and TGFB2R knockout, albeit not a strong one. Importantly, when stressed with TGF-β, only TGFB2R-knockout cells, with or without CARs, retained their cytotoxicity, while CAR-only cells experienced quick loss of function.

Longer survival in vivo

The researchers observed similar results in a mouse model of HCC: TGFB2R knockout seemed to contribute the most toward NK’s anti-cancer activity, with or without CARs, markedly improving survival. NKs that only carried CARs were much less effective.

“These studies demonstrate that it is crucial to block transforming growth factor beta — at least for NK cells, but I also think it’s true for CAR T cells,” said Dan Kaufman, the lead author of this study. “If you unleash NK cells by blocking this inhibitory pathway, they should kill cancer quite nicely. Anyone developing such therapies for solid tumors should be working to inhibit transforming growth factor beta activity to improve cancer-killing and attain effective anti-tumor activity.”

NK cells are generally easier to mass-produce from iPSCs than T cells because the latter require more intricate differentiation protocols and thymus-like conditions to mature. Genetic modifications are also easier to perform in NK cells. NK cells require less personalization and can be used in donor-derived (allogeneic) settings more readily than T cells. As part of the innate immune system, NK cells have a broad range of activity against tumor cells without requiring prior sensitization to specific antigens. This all adds to NK cells’ promise in future anti-cancer therapies [3].

These studies demonstrate that even with NK cells engineered to express tumor-specific CARs, the immunosuppressive activity of TGF-β prevented effective anti-tumor activity if TGF-β activity is not inhibited. To disrupt TGF-β signaling, we utilized 2 different strategies via either deletion of TGFBR2 or over expression of the DN form of TGFBR2 in iPSC-derived NK cells. Both TGFBR2-KO and TGFBR2-DN NK cells demonstrated effective anti-HCC activity even without CAR expression, while CAR-NK cells without inhibition of TGF-β activity had little anti-HCC activity.

Literature

[1] Thangaraj, J. L., Coffey, M., Lopez, E., & Kaufman, D. S. (2024). Disruption of TGF-β signaling pathway is required to mediate effective killing of hepatocellular carcinoma by human iPSC-derived NK cells. Cell Stem Cell.

[2] Baba, A. B., Rah, B., Bhat, G. R., Mushtaq, I., Parveen, S., Hassan, R., … & Afroze, D. (2022). Transforming growth factor-beta (TGF-β) signaling in cancer-A betrayal within. Frontiers in pharmacology, 13, 791272.

[3] Goldenson, B. H., Hor, P., & Kaufman, D. S. (2022). iPSC-derived natural killer cell therapies-expansion and targeting. Frontiers in immunology, 13, 841107.

Researchers have found that men and women have significant differences in how their brains’ blood vessels change in Alzheimer’s disease.

The collapse of the blood-brain barrier

This research begins with a discussion of Alzheimer’s and its two principal symptoms: the well-known plaques that coat the brain, and the tau proteins that aggregate in neurons. However, targeting either of these proteins after they have already been formed has not been effective in preventing neurons from dying or stopping cognitive decline [1]. These researchers hypothesize that such interventions simply occur too late. Therefore, they chose to focus on something that occurs much earlier in the process: the decline of the blood-brain barrier (BBB) [2].

This decline is precipitated by vascular aging, which has been recently reported to be associated with Alzheimer’s disease [3]. Men and women do not experience vascular aging in precisely the same way [4, 5]. In older women, a decrease in estrogen causes vascular decline [6], making them more susceptible to Alzheimer’s [7].

Gene expressions in Alzheimer’s differ by sex

This research began with a gene expression analysis using samples from 428 people, looking for differentially experienced genes between Alzheimer’s patients and people who did not have the disease. There were no significant differences between the sexes in cellular numbers. In all Alzheimer’s patients, gene expressions in the prefrontal cortex that were related to the vascular system were altered, while in other regions, the gene expression differences were largely related to neuronal support.

These prefrontal cortex differences were much more pronounced in men. Further analysis found that they were largely found in endothelial cells, which line blood vessel walls and form the BBB. Many of these gene expression differences had already been noted as being risk factors for Alzheimer’s.

Men and women did share some similarities in how their genes changed expression with Alzheimer’s, most notably in the metabolism of lipids (fats) and in neuroprotective genes. However, many of the endothelial cells’ gene expression differences were not the same in men and women, and often, they were in opposite directions; some of the relevant genes that are commonly upregulated in men with Alzheimer’s were likely to be downregulated in women with the disease and vice versa.

The hypoxic response

One of the most critical gene sets involved the hypoxic response, which occurs when cells don’t get enough oxygen. The downstream pathways involved in hypoxia were activated in men but not women. The researchers were even able to confirm this in model mice; just like in people, male mice with Alzheimer’s activated the hypoxic response, while female mice did not.

One consequence of the hypoxic response is angiogenesis: the creation of new blood vessels. Genes related to the hypoxic response leading to angiogenesis were strongly upregulated in men with Alzheimer’s, but in women, they were not. However, in women, a different gene, PIK3C2A, was upregulated that leads to angiogenesis; the researchers suggest that this may be a druggable target for men, who do not normally upregulate it.

The researchers investigated a core pathway involved in angiogenesis in women. CREB1 affects vascular endothelial growth factor (VEGF), which affects angiogenesis and endothelial cells. They noted previous work showing that estrogen strongly affects this pathway [8] and that the protein CREB has been investigated as a druggable target in Alzheimer’s.

Estrogen also affects CREB1, and these researchers pinpointed the menopause-related decline in estrogen as being key to the onset of Alzheimer’s in women. An estrogen receptor and a receptor for a hypoxia-inducible factor both have many of the same downstream targets, and menopause is a significant risk factor for vascular disease [9]. Therefore, these researchers believe that men retain a key protection in the blood-brain barrier that post-menopausal women do not. Hormone replacement therapy for older women, therefore, may be found to be necessary to prevent late-life Alzheimer’s, or another method of activating the relevant angiogenesis pathways may be found.

Literature

[1] Godyń, J., Jończyk, J., Panek, D., & Malawska, B. (2016). Therapeutic strategies for Alzheimer’s disease in clinical trials. Pharmacological Reports, 68(1), 127-138.

[2] Sweeney, M. D., Zhao, Z., Montagne, A., Nelson, A. R., & Zlokovic, B. V. (2018). Blood-brain barrier: from physiology to disease and back. Physiological reviews.

[3] Oh, H. S. H., Rutledge, J., Nachun, D., Pálovics, R., Abiose, O., Moran-Losada, P., … & Wyss-Coray, T. (2023). Organ aging signatures in the plasma proteome track health and disease. Nature, 624(7990), 164-172.

[4] Ji, H., Kwan, A. C., Chen, M. T., Ouyang, D., Ebinger, J. E., Bell, S. P., … & Cheng, S. (2022). Sex differences in myocardial and vascular aging. Circulation research, 130(4), 566-577.

[5] DuPont, J. J., Kim, S. K., Kenney, R. M., & Jaffe, I. Z. (2021). Sex differences in the time course and mechanisms of vascular and cardiac aging in mice: role of the smooth muscle cell mineralocorticoid receptor. American Journal of Physiology-Heart and Circulatory Physiology, 320(1), H169-H180.

[6] Moreau, K. L., Hildreth, K. L., Meditz, A. L., Deane, K. D., & Kohrt, W. M. (2012). Endothelial function is impaired across the stages of the menopause transition in healthy women. The Journal of Clinical Endocrinology & Metabolism, 97(12), 4692-4700.

[7] Arvanitakis, Z., Capuano, A. W., Leurgans, S. E., Bennett, D. A., & Schneider, J. A. (2016). Relation of cerebral vessel disease to Alzheimer’s disease dementia and cognitive function in elderly people: a cross-sectional study. The Lancet Neurology, 15(9), 934-943.

[8] Burtscher, J., Mallet, R. T., Burtscher, M., & Millet, G. P. (2021). Hypoxia and brain aging: Neurodegeneration or neuroprotection?. Ageing research reviews, 68, 101343.

[9] El Khoudary, S. R., Aggarwal, B., Beckie, T. M., Hodis, H. N., Johnson, A. E., Langer, R. D., … & American Heart Association Prevention Science Committee of the Council on Epidemiology and Prevention; and Council on Cardiovascular and Stroke Nursing. (2020). Menopause transition and cardiovascular disease risk: implications for timing of early prevention: a scientific statement from the American Heart Association. Circulation, 142(25), e506-e532.