Evidence suggesting that high intake
of fluoride provokes nephrolithiasis in tribal populations.
Singh PP, Barjatiya MK, Dhing S, Bhatnagar R, Kothari S, Dhar V.
Department of Biochemistry, Darshan Dental College, Madhuvan, India. firstname.lastname@example.org
The present study was designed to evaluate the role of fluoride in urolithiasis in humans. Two areas were selected for this purpose, a fluoride endemic area (EA) and a fluoride non-endemic area (NEA). The prevalence of urolithiasis was 4.6 times higher in EA than in NEA. Furthermore, the prevalence was almost double in subjects with fluorosis than without fluorosis in the endemic area. No relationship was observed between urolithiasis and the duration of fluorosis. The fluoride levels in drinking water ranged from 3.5 to 4.9 ppm in EA and subjects from this area excreted more fluoride. A comparison of normal subjects (NS) from EA and NEA revealed that endemic subjects tend to have slightly higher mean serum thiobarbituric acid reactive substance (TBAR) levels and excrete more oxalate and fluoride than their non-endemic counterparts. The urinary stone formers (SF) from the two areas showed a similar tendency, though again the difference was not significant. Citrate excretion in SF was almost normal in the EA, but NEA SF had significantly lower excretion levels. Urinary stones from endemic patients had higher fluoride, oxalate and calcium levels than those from non-endemic patients. In vitro studies suggested that fluoride did not influence the heterogonous mineralization of calcium oxalate. In conclusion, the data suggest that fluoride in vivo may behave as a mild promoter of urinary stone formation by (a) excretion of insoluble calcium fluoride, (b) increasing oxalate excretion and (c) mildly increasing the oxidative burden.
Prevalence of urolithiasis is high in many parts of India [10, 17, 23, 35, 39, 42] including Rajasthan [26, 27, 43, 44]. The involvement of possible etiological factors in this disease has been reported from time to time [4, 25, 29, 38, 39, 40, 41, 45]. In an earlier study, no relationship was observed between fluoride in drinking water and the prevalence of urinary stone disease in a NEA (Non-Endemic Area) . However, a recent, preliminary study by our group noted a high prevalence of urolithiasis in EA (Endemic Area). The present, extended study supports this finding and further suggests that chronic fluoride consumption increases the risk of urinary stone formation by altering the urinary profile and mildly increasing the oxidative environment.
A door to door survey for urolithiasis among non-tribal people in this region showed a prevalence of 310/100,000, with a male/female ratio of 5.5:1 . On the other hand, this study, as well as other studies from this region [30, 43, 45], indicate that the prevalence is much lower among tribal people. Environmental provocations are known to enhance nephrolithiasis [4, 11] and Barjatiya and Singh  have already reported higher prevalence of nephrolithiasis among tribal people working in a mining environment in this region. In the current study we observed a 4.6-times higher prevalence in EA than in NEA. The role of fluoride as a urinary stone risk factor is further supported by the finding that prevalence in fluorotic patients was double (1,071/100,000) that found in non-fluorotic patients (578/100,000). However, no relation was found between the duration of fluorosis and the prevalence of urolithiasis. This can be attributed to its multifactorial etiology.
The main contributor to fluoride intake in humans is drinking water and in Rajasthan water fluoride levels in EA are in the range of 2 to 20 ppm [20,50]. In the EA under study, fluoride levels ranged from 3.5 to 4.9 ppm. According to the fluoride map of Rajasthan, the populations of Udaipur and Dungarpur districts suffer from a relatively moderate degree of fluoride toxicity . Since the concentration of urinary constituents is the most important determinant of urinary stone formation and growth, urine chemistry of tribal NS and SF from EA and NEA was examined for stone promoters and inhibitors. In NEA the urinary profile of NS was comparable to other studies [4, 26, 45]. The SF in this population, as in earlier studies [43, 45], showed three important differences from NS, i.e., higher excretion of oxalate and calcium and lower excretion of citrate. However, the level of calcium does not appear to be an important contributing factor. In the EA both NS and SF had either significantly higher oxalate excretion or tended to have higher excretion. Citrate excretion was normal.
Subsequently, NS and SF from EA were compared with their counterparts in NEA. The NS from EA showed two important differences from NS from NEA, i.e., significantly higher oxalate and fluoride and lower magnesium excretion. The SF from these two areas also showed similar trends, though oxalate and magnesium did not attain the level of significance. However, an additional change in SF from EA was the normal level of citrate excretion. Citrate is undoubtedly a good inhibitor of urinary stone formation and plays multiple roles in the prevention of calcium lithiasis . However, its relative potential as an inhibitor is expected to lose much of its importance in this population which has low calcium excretion. The NS in EA showed a tendency to have slightly higher TBAR levels and SF in both groups had slightly higher levels than their NS counterparts. These data, and our unpublished data on animals, suggest that fluoride slightly increases the oxidative environment in situ. Higher oxidative load in SF compared to NS has been reported earlier, also from Rajasthan . An enlarged oxalate pool in vivo can lead to oxidative stress  and this stress can damage or kill the renal epithelial cells  which can then serve as a site or nidus for the formation of a urinary stone.
There are a number of controversies on the behavior of fluoride in the context of lithogenesis:
Why did Teotia et al.  not find any relationship between fluorosis and bladder stone disease in children whereas others have noted a high prevalence in humans and in experimental animals with a high fluoride intake from drinking water The conclusions derived by Teotia et al.  are suspect because they neither studied bladder stone disease nor examined the prevalence and the urine chemistry of NS and SF, in EA. Herring et al  were precipitate in concluding that fluoride is an inhibitor because they examined neither the frequency of calcium oxalate crystals nor the size of crystals nor urinary oxalate which are prerequisites for their conclusion. They also missed the point that fluoride is a highly reactive anion and quickly forms calcium fluoride which is insoluble in water. In all probability, the lower calcium content in the kidneys of their rats receiving high fluoride is due to the lower availability of calcium for absorption. This is because calcium fluoride precipitates in the intestine and would not be available for absorption. For the same reason, there would have been diminished availability of calcium in their in vitro experiments for the formation of calcium oxalate. A close scrutiny reveals that the primary cause for bladder stones in Anasuya's experiment was diet related and not due to fluoride because 25% and 33% of rats respectively developed bladder stones on low and high calcium control diets during the 10-week period of study. Fluoride only accentuated the risk of stone formation..
Why Anasuya  found fluoride to be a promoter in rats whereas Herring et al. , found it to be an inhibitor
Why some in vitro studies found fluoride to be a promoter  whereas others found it to be unrelated to the risk of urinary stone formation, and
Why the present study assigns fluoride the role of a promoter of urinary stones
In our study, the stones from endemic SF have significantly higher fluoride content and calcium content also tended to be higher. On the basis of his observations Machov  concluded that fluoride was a permanent constituent of urinary stones. He also suggested that it is an active constituent. Our data are in conformity with his findings although we do not believe that fluoride plays an active role. Our view is also supported by the work of Anasuya , who found that weights of stones in rats with low and high fluoride intake were similar. Most likely, the insoluble calcium fluoride in water is fortuitously incorporated into the stone.
While Anasuya and Rao  noted fluoride to be a promoter in their in vitro studies, our data do not support their observations. The increased deposition of calcium on tendons in their study may be due to deposition of water insoluble calcium fluoride. We did not find any change in calcium uptake when this possibility was eliminated.
Taking into consideration all of the data from our study and from the studies of others, we believe that the chronic intake of fluoride increases the risk of urinary stone formation. The proposed mechanism of fluoride participation in lithogenic processes in the tribal population studied could be as follows: The Rajasthani population in general, including tribal people, suffer from a high excretion of oxalate, mainly due to defective nutrition [37, 38, 43, 45]. Fluoride may be indirectly increasing oxaluria by enhancing the absorption of oxalate from the intestine due to low availability of calcium as part of the intestinal calcium is precipitated as calcium fluoride. Oxalate in -vitro, as well as in -vivo, increases oxidative load [33, 47] which may injure or kill renal epithelial cells, providing an opportunity for urinary crystals to attach to the injured site or to be deposited on dead epithelial cells which serve as a nidus. Fluoride may also be synergistically acting with oxalate to potentiate the oxidative environment. Fluoride is also thought to possess the inherent property of being able to bind crystals strongly, which gives more hardness to bone . It is, therefore, logical to presume that the formation of a urinary stone will occur when these conditions overwhelm the inhibitory forces present.
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