4. Results
4.6. Organ Weight
In study one, with Normal diet, only some organs, brain, liver and spleen were measured at the necropsy. In study two, with Low energy diet, all important organs were measured. In this case comparisons can be taken in brain, liver and spleen weights, analysing the absolute and relative organ weights referring to the body weight.
The absolute and relative brain weight and spleen weight did not differ
significantly in the male animals fed with different diets. Slight increase of absolute liver weight occurred in male rats fed with Low Energy Diet in spite of the lower body weight. In relative liver weight analysis 26% difference occurred.
The relative liver weight of male animals fed with Low Energy Diet was significantly higher than the relative liver weight of male animals fed with Normal Diet.
In female animals the absolute liver weight was significantly higher in group fed with Low Energy Diet then the mean weight of this organ in the group fed with Normal Diet. The correction with the body weight in analysis of relative liver weight decreased the observed difference of this parameter to 12%, which is on the border of the biological significance.
4.7. Histopathology
At the histological evaluation of the sections of different organs we observed different types of benign and malignant tumours, non-neoplastic hyperplastic lesions and non-neoplastic other lesions (degenerations, necrosis, inflammations etc.)
It should be mentioned that a neoplasm is a new growth of cells that (1) proliferate continuously without control (2) bear a considerable resemblance to the healthy cells from which they arose, (3) have no orderly structural arrangement, (4) serve no useful function and (5) for the present, at least, have no clearly understood cause.
Because of great destructiveness, neoplasms are classed as malignant, while those that grow only expansively, much less dangerous to their host, are said to be benign.
The main characteristics of malignant and benign neoplasms are demonstrated in Table 10.
Table 10 Comparison of Benign and Malignant Neoplasms
Characteristic Benign Malignant
Growth rate Slow Rapid
Growth limits Circumscribed Unrestricted
Mode of growth Expansion Invasion
Differentiation Good Anaplastic
Stroma Usually abundant Usually scant
Metastasis None Frequent
Recurrence Rare Frequent
The observed types of benign tumours were the follows:
Table 11 Benign Neoplasms
Organs Normal Diet Low Energy Diet
Hypophysis adenoma* adenoma* TUMOUR in the thoracic or
abdominal cavity -- fibroma
Liver -- cholangioma
cortical adenoma cortical adenoma Adrenal gland
pheochromocytoma pheochromocytoma Thyroid gland follicular adenoma follicular adenoma
Pancreas -- adenoma
Spleen -- haemangioma
seminoma seminoma Testis
Leydig-cell tumour Leydig-cell tumour
adenoma adenoma
leiomyoma -- Uterus
myxoma myxoma REMARK:
∗ = It should be noted that some descriptions consider the hypophysis adenoma as a benign type of tumour and incidental lesion. In this experiment the adenomas were often large and compressed the normal tissue of adenohypophysis and caused death of animals.
The observed types of malignant tumours were the follows:
Table 12 Malignant Neoplasms
Organ Normal Diet Low Energy Diet
adenocarcinoma adenocarcinoma
carcinoma (undifferentiated) carcinoma (undifferentiated)
Liver -- hepatocellular carcinoma
Intestine fibrosarcoma --
Lung adenocarcinoma --
Kidneys -- adenocarcinoma
Heart sarcoma --
Thyroid gland follicular carcinoma --
Brain astrocytoma astrocytoma
Prostate fibrosarcoma --
Bone -- osteosarcoma
Eye -- carcinoma planocellulare
Table 13 Non-neoplastic hyperplastic lesions
Organ Normal Diet Low Energy Diet
hyperplastic nodule hyperplastic nodule Liver
bile duct hyperplasia bile duct hyperplasia
Thyroid gland -- C-cell hyperplasia
Pancreas islet-cell hyperplasia islet-cell hyperplasia
Table 14 Non-neoplastic other lesions
Organ Normal Diet Low Energy Diet
Hypophysis cyst cyst
-- atheroma Skin
inflammation --
-- vacuolisation of hepatocytes
fatty infiltration -- Liver
-- coagulative necrosis
Kidney chronic nephropathy chronic nephropathy
Stomach -- ulcer
oedema oedema Lung
-- bronchopneumonia
Heart fibrosis fibrosis
Adrenal gland cystic degeneration cystic degeneration The number of histologically investigated animals were 200 and of investigated organs/tissue samples were about 10.000.
In Group 1 fed with normal diet the most common causes of death were:
hypophysis adenoma, malignant (often generalized) tumours, chronic nephropathy, lung oedema and serious cystic degeneration of the adrenal glands.
In Group 2 fed with low energy diet, similarly to the Normal diet group, the most common causes of death were: hypophysis adenoma, malignant tumours, chronic nephropathy, lung oedema and cystic degeneration of the adrenal glands
Hypophysis adenoma
Hypophysis adenomas are well-delineated masses that compress the surrounding parenchyma. The neoplastic cells are arranged in solid sheets or compact branching cords. The vascular pattern may be more or less evident, but large neoplasms often have angiectasis, cyst-like lesions and/or haemorrhage.
According to the historical literature data the hypophysis adenomas are common in Charles River CD rats. The incidence varies from 0 % to 90 % in old male and female animals.
In this study - according to the historical data - hypophysis adenomas were the most common tumours. See Figure 13.
Figure 13 Photo of hypophysis adenoma
Other malignant tumours
Undifferentiated sarcoma in the mammary gland, heart, vagina, undifferentiated carcinoma in the skin, vagina or the abdominal cavity, myxosarcoma in the skin, uterus or in the abdominal cavity, follicular carcinoma in the thyroid gland, plexus carcinoma and astrocytoma in the brain, osteosarcoma in the bone, carcinoma planocellulare in the eye were detectable only sporadically.
The malignant tumours which caused death of animals were the sarcoma polymorphocellulare, fibrosarcoma, lymphoma malignum, adenocarcinoma, and hepatocellular carcinoma, which were infrequent in both males and females.
Benign tumours
The most common benign tumour was in this study the mammary gland adenoma in the female animals.
The other types of benign neoplasms (fibroma in the mammary gland, skin, thoracic or abdominal cavity, myxoma, fibro-adenoma, adeno-papilloma in the skin, adenoma in the kidney, adrenal gland, thyroid gland, parathyroid gland, salivary gland, pancreas, uterus, lipoma in the wall of stomach, thymoma in the thymus, pheochromocytoma in the adrenal gland, myxoma, fibro-papilloma, leiomyoma in the uterus, haemangioma in the spleen and lymph node) had generally low frequency and had the same incidence in both groups.
Non-neoplastic hyperplastic lesions
At the histological evaluation of different organs of experimental animals we observed non neoplastic hyperplastic lesions (hyperplastic nodules and bile duct hyperplasia in the liver, C-cell hyperplasia in the thyroid gland, islet-cell hyperplasia in the pancreas, cystic hyperplasia in the seminal vesicle) as well.
Hyperplastic nodules in the liver were rare and occurred in the female animals fed with low energy diet.
The foci of cellular alteration belonged mainly to hyperplasias, that are perceived to be secondary, non-neoplastic responses to degenerative changes of the liver.
Some focus of cellular alteration was basophilic, and eosinophilic character. No compression of surrounding parenchyma or higher incidence of mitotic figures were seen.
Bile duct hyperplasia is a common aging lesion in rats often accompanied by perilobulary fibrosis.
The C-cell hyperplasia in the thyroid gland the cystic hyperplasia in the seminal vesicle and the islet-cell hyperplasia in the pancreas were detectable only sporadically.
Table 15 includes the overall incidence of benign and malignant tumours and hyperplastic findings by groups and sex at the fatal death and moribund condition and at the terminal sacrifice.
Table 15 Overall Incidence of Benign and Malignant Tumours
Fatal Death and Moribund Terminal Sacrifice Normal Diet Low Energy Diet Normal Diet Low Energy Diet LESIONS
M F M F M F M F
No. of animals 22 23 17 24a 28 27 33 25
bening tumours 10 20 14 35 21 33 32 26
malignant tumours 6 10 7 7 4 3 3 2
tumours (total) 16 30 21 42 25 36 35 28
hypophysis adenoma 7 13 10 21 5 18 19 10
non-neoplastic
hyperplasias 4 2 5 9 13 6 17 8
hypophysis
adenoma/animal 0,32 0,57 0,59 0,88 0,18 0,67 0,58 0,40 bening tumours / animal 0,45 0,87 0,82 1,46 0,75 1,22 0,97 1,04 malignant
tumours/animal 0,27 0,43 0,41 0,29 0,14 0,11 0,09 0,08 tumours (total) / animal 0,73 1,30 1,24 1,75 0,89 1,33 1,06 1,12
non-neoplastic
hyperplasias / animal 0,18 0,09 0,29 0,38 0,46 0,22 0,52 0,32 Remark:
a = one animal was not evaluated due to autolysis.
Considering the results summarised in Table 15, the total number of tumours are higher in male and female animals fed with Low Energy Diet, than in the male and female groups fed with Normal Diet. It can improve the sensitivity of statistical analysis and helps to detect true test-item effect in bioassays.