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Pang, Li Yin (2024) Exploring the effects of medicinal fungi on isolated bladder strips (ex vivo) and Caenorhabditis elegans (in vivo): focus on the caterpillar fungus, Ophiocordyceps sinensis. PhD thesis, University of Nottingham.

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Abstract

Medicinal fungi have been traditionally used to treat many medical conditions. Today, they remain a major source of new pharmaceutical products due to the presence of a plethora of bioactive components. In China and several East Asian countries, Ophiocordyceps sinensis, one of the most sought-after medicinal fungi, is widely used for alleviation of urinary symptoms such as frequent urination and nocturia – the two most common symptoms of overactive bladder syndrome (OAB). Despite this, there is no credible scientific evidence that links the effect of O. sinensis to bladder function. Therefore, this study was initiated to explore the therapeutic potential of medicinal fungi in OAB due to the limitations of the current conventional treatment options, and to provide scientific validation of O. sinensis in alleviating OAB symptoms. Specifically, the principal aim of this study was to investigate the pharmacological actions of O. sinensis extract in bladder by using the isolated tissue bath technique. The secondary aim was to provide evidence on its health benefits, given its established antioxidant properties and that OAB is highly prevalent in the geriatric population. A cultivated strain of O. sinensis (OCS02®) that has been authenticated by genetic identification was utilised in this study.

In bladder strips pre-contracted with carbachol (muscarinic receptor agonist), the hot water extract of O. sinensis (OCS02-HWE) produced a transient contraction whereas the cold water extract of O. sinensis (OCS02-CWE) elicited a biphasic response that consists of a transient contraction followed by a sustained relaxation response. To elucidate the bioactive components responsible for the bladder relaxant effect, OCS02-CWE was fractionated using size-exclusion chromatography. All the fractions exhibited different effects on the carbachol-pre-contracted bladder strips in which the high molecular weight fraction (OCS02-HMW) demonstrated a weak but sustained relaxation response; the medium molecular weight fraction (OCS02-MMW) produced a marked but non-sustained relaxation response; the low molecular weight fraction (OCS02-LMW) elicited only a transient contractile response, like OCS02-HWE. Based on these results as well as their respective carbohydrates and protein contents, we postulated that the contractile effect produced by O. sinensis could be attributed to heat-stable, carbohydrate components of low molecular weight. In contrast, the potent and sustained relaxant effect could be contributed by heat-labile components that present in both HMW and MMW fractions. To ensure that the maximum relaxation response of O. sinensis is achieved, OCS02-CWE was used for subsequent investigations.

Through a series of mechanistic studies and the use of pharmacological tools, it appeared that the contractile and relaxant effects of OCS02-CWE were mediated via different pathways and did not involve muscarinic receptors, purinergic receptors, adrenoceptors, 5-hydroxytrptamine receptors and cyclic nucleotides. The relaxant effect of OCS02-CWE is partly urothelium-dependent, likely to be mediated via production of NO and regulation of intracellular Ca2+. This is evidenced by the attenuation of OCS02-CWE-induced relaxation response following the removal of urothelium and in the presence of L-NAME (nitric oxide synthase inhibitor) or thapsigargin (sarcoplasmic/endoplasmic reticulum Ca2+-ATPase pump inhibitor). In those conditions, the transient contraction induced by OCS02-CWE was not significantly altered. It was nevertheless absent in KCl-pre-contracted bladder strips, suggesting that the transient contraction was a result of membrane depolarisation. Moreover, the transient contraction was potentiated by TEA (non-selective K+ channel blocker) and attenuated by membrane permeable Ca2+ blockers including nifedipine, verapamil, SKF-96365 and schwarzinicine A. TEA is known to increase the duration and amplitude of an action potential, and membrane depolarisation leads to Ca2+ influx via membrane permeable Ca2+ channels. The transient contraction of OCS02-CWE was also attenuated in the presence of the cold water extract of Lignosus rhinocerus (TM02-CWE), a medicinal fungus that exhibits potent Ca2+ inhibitory activity in smooth muscle.

To offer an insight into the use of medicinal fungi to treat OAB, we assessed the combined effect of OCS02-CWE and TM02-CWE in bladder relaxation. Through response-additive model, we showed that these extracts produced synergistic bladder relaxant effect when they are used at equal proportion and at concentrations below 4 mg/mL. They also appeared to extend lifespan and promote health in Caenorhabditis elegans. In summary, medicinal fungi contain bioactive components that could be further developed to treat OAB and improve health. For this purpose, O. sinensis that exhibits bladder relaxant effect via the regulation of NO and Ca2+ represents a promising source that is worthy of further investigation.

Item Type:	Thesis (University of Nottingham only) (PhD)
Supervisors:	Ting, Kang Nee Lim, Kuan Hon Then, Sue-Mian
Keywords:	Ophiocordyceps sinensis, overactive bladder syndrome (OAB), medicinal fungi, bladder relaxation, Caenorhabditis elegans
Subjects:	Q Science > QD Chemistry > QD241 Organic chemistry
Faculties/Schools:	University of Nottingham, Malaysia > Faculty of Science and Engineering — Science > School of Pharmacy
Item ID:	74375
Depositing User:	Pang, Li
Date Deposited:	09 Mar 2024 04:40
Last Modified:	11 Mar 2024 02:40
URI:	https://eprints.nottingham.ac.uk/id/eprint/74375

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