<div>Anosmia and dysgeusia among COVID-19 patients are associated with low
levels of serum Glucagon-like peptide 1 (GLP-1)</div><div>Abstract</div><div>Purpose: Anosmia and dysgeusia (AD) are common among COVID-19 patients.
These symptoms are not frequently associated with rhinorrhea or nasal
congestion and the underlying mechanism is unclear. Previous reports
suggested that Glucagon-like peptide-1 (GLP-1) signaling plays a role in
the modulation of olfaction and geusia. We aimed to assess the
correlation between GLP-1 and COVID-19-associated AD.</div><div>Methods: Blood samples obtained from COVID-19 patients with and without
AD were tested for serum GLP-1 levels using enzyme-linked immunosorbent
assay (ELISA). A second control group comprised of COVID-19-negative
volunteers.</div><div>Results: Forty-nine subjects were included in the study. Nineteen were
positive for COVID-19. Of the 19 patients, ten had AD and nine declined
such complaints. Age and basic metabolic rate were similar among all
study groups. Serum GLP-1 levels were significantly lower among patients
with AD as compared with patients without AD and COVID-19-negative
individuals (1820 pg/ml vs 3536 pg/ml vs 3014 pg/ml, respectively,
p&lt;0.02).</div><div>Conclusion: COVID-19 patients who reported of AD had lower serum levels
of GLP-1 as compared with those lacking AD symptoms and
COVID-19-negative individuals. These results suggest that GLP-1 may be
involved in the pathogenesis of AD. However, further larger scale
studies should corroborate our findings</div><div>Key words: Anosmia; dysgeusia, COVID-19; Glucagon-like peptide-1</div><div>Introduction</div><div>Olfactory and taste disorders are known to be related to a wide range of
viral infections including SARS-CoV-2. However, the pathophysiological
mechanism has yet to be determined. Many viruses lead to anosmia and/or
dysgeusia (AD) through an inflammatory reaction of the nasal mucosa
leading to rhinorrhea and nasal congestion. However, among COVID-19
patients, especially of mild severity, AD may present as an isolated
complaint (1). In a recent study, of 2013
COVID-19 patients, 1754 patients (87%) reported loss of smell, whereas
1136 (56%) reported taste dysfunction. Both complaints were more common
among mild COVID-19 patients (2). Angiotensin-converting enzyme-2 (ACE2)
is the main host cell receptor that mediates SARS-CoV-2 invasion to
cells, and is widely expressed in multiple tissues including the
epithelial cells of the oral mucosa. In fact, this receptor was highly
enriched in epithelial cells of tongue (3). An inflammatory response
mediated by cytokines leading to oral and olfactory mucosal injury was
suggested as a possible mechanism for AD
(4).</div><div>Interestingly, the signaling pathway of taste modulation and olfaction
involves several ligands, among them is glucagon-like peptide-1 (GLP-1)
(5). GLP-1 is a pleiotropic gut hormone with numerous metabolic
functions and has a main role in glucose homeostasis. GLP-1 is released
in the gut in response to nutrient ingestion and its activity is
regulated by the enzyme dipetidyl-peptidase-4. The taste cells,
particularly L-cells, produce GLP-1 among other hormones
(6). Several reports suggest that GLP-1
functions as a neurotransmitter mediating taste and olfaction (5, 7, 8).
Mice lacking the GLP-1 receptor have significantly reduced taste
sensitivity to sweeteners (7). Pre-pro-glucagon (the precursor of GLP-1)
and GLP-1 receptor mRNAs have been identified in cell layers in the
olfactory bulb suggesting that GLP-1 may mediate information flow from
the olfactory epithelium to the brain (8).</div><div>In view of its role in modulation of gustatory and olfaction functions,
we aimed to examine the association between serum GLP-1 levels among
COVID-19 patients reporting AD versus patients and COVID-19-negative
individuals with intact olfaction and taste.</div><div>Materials and Methods</div><div>The study took place in the COVID-19 wards of Shaare Zedek Medical
center (SZMC), Jerusalem, Israel. We established a new bio-bank of
various blood-derived products from hospitalized COVID-19 patients.
COVID-19 was confirmed based on PCR positive nasopharyngeal swab
specimen tested for SARS-CoV-2. Our study included COVID-19 confirmed
patients with AD symptoms (group A), without AD symptoms (group B) as
well as COVID-19-negative asymptomatic subjects that served as a second
control group (group C). Clinical and demographic data was retrieved
from the computerized medical records or by interviewing the study
participants. Those included: age, sex, comorbidities, presence of
anosmia/dysosmia and or/ dysgeusia and patients’ medications. Severity
of COVID-19 was assessed using the NIH guidelines
(https://www.covid19treatmentguidelines.nih.gov).</div><div>Blood samples were collected from COVID-19-confirmed patients (based on
nasopharyngeal PCR testing) and from COVID-19-negative individuals in 5
mL CAT tubes (456018, Vacuette; Greiner Bio-one). Samples were
centrifuged at 1800 x g, R/T for 20 minutes according to protocol, sera
were separated and aliquots were stored in a dedicated
-80<sup>o</sup>c COVID-19 freezer. Serum GLP-1 levels were
measured using a commercial enzyme-linked immunosorbent assay (ELISA)
according to the manufacturer instructions (abcam, Cambridge, UK).
Differences between groups were assessed using Chi-square test for
categorical variables and Welch analysis of variance (ANOVA) test for
continuous variables with skewed distribution. P value &lt;0.05
was considered statistically significant. Informed consent was obtained
from all participants. The study was approved by the Institutional
Review Board of SZMC.</div><div>Results</div><div>Forty-nine subjects were included in the study. Nineteen were
COVID-19-confirmed hospitalized patients. Of them, 11 (57.9%) had mild
disease, 3 (15.8%) had moderate disease and 5 (26.3%) had severe
illness. One patient with severe COVID-19 died. Overall, 9 COVID-19
patients had complained of AD (group A) and 10 had no AD symptoms (group
B). A third group of 30 asymptomatic COVID-19 negative subjects (group
C) served as a second control group. The demographic and clinical
characteristics of all subjects are shown in Table. None of the patients
with AD had rhinitis or rhinorrhea.</div><div>Age and BMI did not significantly differ between groups. The median age
of patients in groups A, B and C was 52 (range 34-62), 64 (range 40-80)
and 60 years (range 40-69), respectively (p=0.06); with male
predominance (57.1%) in all groups. Height and weight data was missing
in 5 subjects. COVID-19 patients were more obese than COVID-19 negative
controls (Table).</div><div>Type 2 diabetes mellitus was reported in 22.2% (2/9) of patients in
group A, 40% (4/10) in group B and 3.3% (1/30) in group C (Table). One
diabetic patient with intact olfaction and geusia was treated with GLP-1
receptor analogue (liraglutide). Prevalence of co-morbidities aside of
diabetes (including primarily hypertension, hypothyroidism, and
hyperlipidemia) was similar among Groups A and B (Table).</div><div>The mean serum GLP-1 levels among group A patients were significantly
lower as compared with GLP-1 levels in groups B and C (1820±226 pg/ml
vs. 3536 ±449 pg/ml and 3014±161 pg/ml, p&lt;0.002, respectively)
(Figure 1). Serum GLP-1 levels among groups B and C were similar
(p=0.65). GLP-1 levels were also found significantly low among Group A
patients vs Groups B and C after adjustment to age, sex, diabetes, and
BMI (p=0.02). No correlation was found between severity of disease and
serum GLP-1 levels.</div><div>Discussion</div><div>Among all symptoms arise from COVID-19 infection the changes in taste
and smell, although not life-threatening, seem the most bothersome to
many patients. However, the pathophysiology of anosmia and dysgeusia is
not fully elucidated. In this study we examined the association of serum
GLP-1 levels and AD among COVID-19 patients. GLP-1 levels were
significantly lower among COVID-19 patients with AD as compared with
patients without AD and COVID-19-negative subjects.</div><div>Prevalence of smell and/or taste impairment is high among COVID-19
patients (2, 9-11). A proposed mechanism for this chemo-sensitive
dysfunction was local inflammation and cytokine release in the oral and
olfactory mucosa induced by SARS-CoV-2 tissue invasion (12). The high
expression of ACE2 in the oral mucosa, especially the tongue and taste
buds, as well as the nasal cavity olfactory epithelium supports this
notion (3, 13). However, this theory is questionable in mild COVID-19
cases, where the inflammatory response is less robust and rhinorrhea or
nasal congestion are absent (14). Indeed, these symptoms were not
commonly reported in our cohort.</div><div>Others suggested that the interaction of SARS-CoV-2 with sialic acid
receptors on taste buds, impairs vital components of the salivary mucin.
Reduced sialic acid in the saliva may accelerate degradation of the
gustatory particles (12, 15). However, this does not explain dysosmia.</div><div>Our findings may point to another intriguing mechanism of AD, which may
be related to the gut hormone GLP-1 (5). Aside of its major role as an
incretin, enhancing insulin secretion and maintaining energy
homeostasis, GLP-1 has been reported to carry out protective and
regulatory effects in numerous tissues, including heart, adipose,
muscles, bones, kidneys, liver, lungs, and brain as well as the
olfactory epithelium and the taste buds on the tongue (16, 17). In fact,
it functions as a neurotransmitter, coordinating communication between
the peripheral organs and the brain (5, 18, 19). GLP-1 modulates taste
sensitivity and affects taste preferences
(5, 18, 19). Expression of GLP-1 was
reported in two populations of taste cells (Type II and III cells), and
its receptor has been identified in taste nerve fibers in mice and
rodents (18). GLP-1 producing cells and GLP-1 receptors have also been
found in the olfactory bulb (20).</div><div>In view of the above literature and in-line with our findings, we
propose that low serum levels of GLP-1 may contribute to
anosmia/hyposmia and dysgeusia
among COVD-19 patients.</div><div>No previous reports have examined the association between GLP-1 and
olfaction and geusia during viral infections, and particularly among
COVID-19 patients.</div><div>The decrease in GLP-1 may be explained either by increased degradation
or decreased production of the hormone due to local inflammation in the
oral and nasal cavities. Further research at the tissue and cellular
level may shed light on this notion.</div><div>Our study has limitations. First, the retrospective design has inherent
limitations including recall bias. Second, no histopathological analyses
of samples from the oral and nasal cavity were performed. Third, due to
the small sample size, our findings may not be generalizable, and
subgroup analyses (based on BMI, diabetes, age) was not attainable. This
may had affected our findings. However, it is unclear whether GLP-1
secretion (either basal or postprandial) is decreased or enhanced under
an obese, or diabetic status (21). Importantly, BMI was similar among
the study groups and only one patient in the COVID-19-negative group was
treated with GLP-1 analogue.</div><div>It would be of interest to examine the degree of AD among diabetic
COVID-19 patients who are treated with GLP-1 analogues or among patients
post-bariatric surgery. Both groups have relatively high GLP-1 levels.</div><div>In summary, our preliminary findings point to a possible mechanism by
which decreased GLP-1 may be involved in impairment of olfaction and
geusia among COVID-19 patients. Further research is required to conclude
a causal association.</div><div>Declarations:</div><div>Funding – the study was funded by Shaare Zedek Scientific Ltd.
(Mada’it)</div><div>Conflicts of interest - none to declare</div><div>Availability of data and material - raw data will be uploaded to
Research Square</div><div>Code availability – not applicable</div><div>Ethics approval – the study was approved by the SZMC institutional
board, approval number SZMC-02-0278</div><div>Consent to participate – all study patients signed an informed consent
permitting collection of blood samples for research purposes.</div><div>Consent for publication – all authors permit publication of the study
by the publisher</div><div>References</div><div>1. Lechien JR, Chiesa-Estomba CM, De
Siati DR, Horoi M, Le Bon SD, Rodriguez A et al. Olfactory and gustatory
dysfunctions as a clinical presentation of mild-to-moderate forms of the
coronavirus disease (COVID-19): a multicenter European study. Eur Arch
Otorhinolaryngol. 2020 ;277:2251-2261.</div><div>2. Lechien JR, Chiesa-Estomba CM, Hans
S, Barillari MR, Jouffe L, Saussez S. Loss of Smell and Taste in 2013
European Patients With Mild to Moderate COVID-19. Ann Intern Med. 2020
;173:672-675.</div><div>3. Xu H, Zhong L, Deng J, Peng J, Dan
H, Zeng X, Li T, Chen Q. High expression of ACE2 receptor of 2019-nCoV
on the epithelial cells of oral mucosa. Int J Oral Sci. 2020 ;12:8.</div><div>4. Lozada-Nur F, Chainani-Wu N, Fortuna G, Sroussi H. Dysgeusia in
COVID-19: Possible Mechanisms and Implications. Oral Surg Oral Med Oral
Pathol Oral Radiol. 2020 ;130:344-346.</div><div></div><div>5. Martin B, Maudsley S, White CM, Egan JM. Hormones in the
naso-oropharynx: endocrine modulation of taste and smell. Trends
Endocrinol Metab. 2009 ;20:163-70.</div><div></div><div>6. Müller TD, Finan B, Bloom SR, D’Alessio D, Drucker DJ, Flatt PR et
al. Glucagon-like peptide 1 (GLP-1). Mol Metab. 2019 ;30:72-130.</div><div>7. Martin B, Dotson CD, Shin YK, Ji S, Drucker DJ, Maudsley S, Munger
SD. Modulation of taste sensitivity by GLP-1 signaling in taste buds.
Ann N Y Acad Sci. 2009 ;1170:98-101.</div><div>8. Merchenthaler I, Lane M, Shughrue P. Distribution of pre-pro-glucagon
and glucagon-like peptide-1 receptor messenger RNAs in the rat central
nervous system. J Comp Neurol. 1999 ;403:261-80.</div><div></div><div>9. Aziz M, Perisetti A, Lee-Smith WM, Gajendran M, Bansal P, Goyal H.
Taste Changes (Dysgeusia) in COVID-19: A Systematic Review and
Meta-analysis. Gastroenterology. 2020;159:1132-1133.</div><div></div><div>10. Spinato G, Fabbris C, Polesel J, Cazzador D, Borsetto D, Hopkins C,
Boscolo-Rizzo P. Alterations in Smell or Taste in Mildly Symptomatic
Outpatients With SARS-CoV-2 Infection. JAMA. 2020 ;323:2089-2090.</div><div>11. Speth MM, Singer-Cornelius T, Oberle M, Gengler I, Brockmeier SJ,
Sedaghat AR. Olfactory Dysfunction and Sinonasal Symptomatology in
COVID-19: Prevalence, Severity, Timing, and Associated Characteristics.
Otolaryngol Head Neck Surg. 2020;163:114-120.</div><div>12. Vaira LA, Salzano G, Fois AG,
Piombino P, De Riu G. Potential pathogenesis of ageusia and anosmia in
COVID-19 patients. Int Forum Allergy Rhinol. 2020 ;10:1103-1104.</div><div>13. Butowt R, Bilinska K. SARS-CoV-2: Olfaction, Brain Infection, and
the Urgent Need for Clinical Samples Allowing Earlier Virus
Detection. <i>ACS Chem Neurosci</i> . 2020;11:1200-1203.</div><div>14. Dev N, Sankar J, Gupta N, Meena RC, Singh C, Gupta DK, Sen MK.
COVID-19 with and without anosmia or dysgeusia: A case-control study. J
Med Virol. 2021;93:2499-2504.</div><div></div><div>15. Mirabelli M, Chiefari E, Puccio L, Foti DP, Brunetti A. Potential
Benefits and Harms of Novel Antidiabetic Drugs During COVID-19 Crisis.
Int J Environ Res Public Health. 2020;17:3664.</div><div>16. Muscogiuri G, Cignarelli A, Giorgino F, Prodam F, Santi D, Tirabassi
G, Balercia G, Modica R, Faggiano A, Colao A. GLP-1: benefits beyond
pancreas. J Endocrinol Invest. 2014;37: 1143-53.</div><div>17. Thiebaud N, Llewellyn-Smith IJ, Gribble F, Reimann F, Trapp S,
Fadool DA. The incretin hormone glucagon-like peptide 1 increases mitral
cell excitability by decreasing conductance of a voltage-dependent
potassium channel. J Physiol. 2016;594: 2607-28.</div><div>18. Takai S, Yasumatsu K, Inoue M, Iwata S, Yoshida R, Shigemura N et
al. Glucagon-like peptide-1 is specifically involved in sweet taste
transmission. FASEB J. 2015 ;29:2268-80.</div><div>19. Bagnoli E, FitzGerald U. Mitral cells and the glucagon-like peptide
1 receptor:</div><div>The sweet smell of success? Eur J Neurosci. 2019;49:422-439.</div><div>20. Fadool DA, Tucker K, Pedarzani P. Mitral cells of the olfactory bulb
perform metabolic sensing and are disrupted by obesity at the level of
the Kv1.3 ion channel. PLoS One. 2011;6:e24921.</div><div>21. Hira T, Pinyo J, Hara H. What is GLP-1 Really Doing in Obesity?
Trends Endocrinol Metab. 2020;31:71-80</div>